C. elegans neuromuscular junctions, Drosophila neuromuscular junctions, lamprey reticulospinal giant axons, goldfish retinal ribbon synapses, the calyx of Held, and rodent hippocampal synapses. We will compare these systems in terms of their known modes and kinetics of synaptic vesicle endocytosis, as well as the underlying molecular machineries. We will also provide the future development of this field.Neurotransmission in complex animals depends on a choir of functionally distinct synapses releasing neurotransmitters in a highly coordinated manner. During synaptic signaling, vesicles fuse with the plasma membrane to release their contents. The rate of vesicle fusion is high and can exceed the rate at which synaptic vesicles can be re-supplied by distant sources. Thus, local compensatory endocytosis is needed to replenish the synaptic vesicle pools. Over the last four decades, various experimental methods and model systems have been used to study the cellular and molecular mechanisms underlying synaptic vesicle cycle. Clathrin-mediated endocytosis is thought to be the predominant mechanism for synaptic vesicle recycling. However, recent studies suggest significant contribution from other modes of endocytosis, including fast compensatory endocytosis, activity-dependent bulk endocytosis, ultrafast endocytosis, as well as kiss-and-run. Currently, it is not clear whether a universal model of vesicle recycling exist for all types of synapses. It is possible that each synapse type employs a particular mode of endocytosis. Alternatively, multiple modes of endocytosis operate at the same synapse, and the synapse toggles between different modes depending on its activity level. Here we compile review and research articles based on well-characterized model systems: frog neuromuscular junctions,  These studies led to the discovery of five distinguishable mechanisms: clathrin-mediated endocytosis, fast compensatory endocytosis, activity-dependent bulk endocytosis, ultrafast endocytosis, and kiss-and-run  neuromuscular junctions, Drosophila neuromuscular junctions, lamprey reticulospinal giant axons, ribbon synapses of goldfish retinal bipolar neurons, the calyx of Held, and rodent hippocampal synapses. For each synapse, we will first describe its anatomical and functional features, then review what is known about endocytosis and its molecular requirements in each system. Furthermore, we will discuss whether some earlier observations may be explained by ultrafast endocytosis, since this pathway was only discovered in 2013. We will close with perspectives on the future development of this field.What factors determine the mode of endocytosis at a particular synapse? At any given presynaptic terminal, one form of endocytosis might predominate. Alternatively, multiple forms of endocytosis might cooperate, depending on the availability of vesicles at the terminals and the activity level, to meet the demand for synaptic vesicle recycling. Here, we review the literature concerning synaptic vesicle endocytosis in a number of well-characterized model systems: frog neuromuscular junctions, Rana pipiens) were the model systems used to study synaptic vesicle dynamics. The anatomical and functional features of these neuromuscular junctions have been well characterized since initial work by Katz and colleagues in the 1950s  of 23 s  have not been explored in frog neuromuscular junctions. What little data are available come from studies using chemical probes. For example, the actin polymerization blocker cytochalasin-D prevents the initiation of activity-dependent bulk endocytosis  is an excellent model system for cell biological studies. Adult animals are roughly 1 mm in length and are composed of about 1000 somatic cells, of which exactly 302 are neurons . Finally, despite initial technical challenges, protocols for performing whole-cell patch clamping of motor neurons and the muscles of C. elegans are now well-developed , displays severe locomotion defects  displays large vesicles linked to the plasma membrane by narrow necks close to the dense projection 1 s after a single stimulation pulse, indicating a defect in ultrafast endocytosis  and an accumulation of endosomes  and synaptojanin (unc-26) block synaptic recycling in C. elegans neuromuscular junctions, clathrin itself may not be involved in synaptic vesicle recycling. In mutants with a temperature-sensitive allele of clathrin heavy chain (CHC), shifting to the non-permissive temperature removes almost all CHC but does not significantly reduce the number of synaptic vesicles at steady state or the amplitude of postsynaptic miniature currents  is an excellent model for studying the synaptic development and function  mutation , with both coated and uncoated endocytic intermediates stalled throughout the plasma membrane, although the number of coated pits was noted to be low , which are hubs for Synaptotagmin-1 (Syt I) and AP2, leads to accumulation of fully internalized large endocytic vesicles and a reduced density of synaptic vesicles  and intersectin-1/Dap60  can reach up to 80 μm in diameter. These axons are specialized in transmitting phasic signals by firing brief high-frequency spike trains  and electrical synapses on the dendrites of spinal neurons in the lateral column  but resumes upon re-exposure to Ca2+  is organized in a donut-shape at lamprey reticulospinal synapses, presumably surrounding the active zone . As in retinal bipolar neurons  also emerges with long high-frequency AP trains  is also observed upon repeated RRP-depleting pulses  is also found at hippocampal mossy fiber synapses following a single action potential  following mild stimulation  at room temperature, but only partially blocks recycling processes triggered by stronger stimulation  as separate mechanisms based on their kinetics and molecular requirements, but the most important emerging concept in the field is that unlike the classical view, synaptic vesicle endocytosis is clathrin-independent. Thus, future investigation is warranted to test whether this concept is valid and if so, tease out why clathrin does not function at the plasma membrane under the physiological conditions and under what circumstances is clathrin required at the plasma membrane. In addition, whether all three clathrin-independent endocytic pathways share general mechanisms must be determined.In order to determine the mechanism of synaptic vesicle endocytosis, a combination of multiple experimental approaches is needed, since each approach has distinct strengths and weaknesses. For example, the time-resolved electron microscopy approach is sensitive to events faster than a second, but less sensitive to slow events. In addition, only a single time point can be observed for a given sample due to the nature of electron microscopy, and thus many synapses must be reconstructed (Watanabe, Drosophila neuromuscular junctions and lamprey reticulospinal giant synapses (Takei et al., C. elegans neuromuscular junctions (Pant et al., C. elegans neuromuscular junctions rely less on clathrin-mediated endocytosis than the other two systems. Thus, differences in molecular requirements among model systems is likely due to their differential preference for modes of endocytosis and vesicle regeneration.The molecular requirements of synaptic vesicle recycling in different model systems are, as expected, similar. Dynamin, for example, is almost invariably required for the major endocytic pathways at all seven types of synapses, the only exceptions are dynamin-independent forms of endocytosis found at the calyx of Held and cultured cortical neurons during highly intense stimulation (Xu et al., Finally, the potential diversity of synaptic vesicle endocytosis cannot be fully encompassed by the model systems reviewed here. All seven model systems reviewed here represent classic fast neurotransmission. However, synaptic vesicle endocytosis at slow-releasing synapses might use different mechanisms. For instance, kiss-and-run, which is not prominently featured in the model systems reviewed here, is a major mechanism underlying exo/endocytosis in chromaffin cells (Perrais et al., QG and SW contributed equally to the work.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.


Progress in iPSC-based cellular systems provides new insights into human brain development and early neurodevelopmental deviations in psychiatric disorders. Among these, studies on schizophrenia (SCZ) take a prominent role owing to its high heritability and multifarious evidence that it evolves from a genetically induced vulnerability in brain development. Recent iPSC studies on patients with SCZ indicate that functional impairments of neural progenitor cells (NPCs) in monolayer culture extend to brain organoids by disrupting neocorticogenesis in an in vitro model. In addition, the formation of hippocampal circuit-like structures in vitro is impaired in patients with SCZ as is the case for glia development. Intriguingly, chimeric-mice experiments show altered oligodendrocyte and astrocyte development in vivo that highlights the importance of cell–cell interactions in the pathogenesis of early-onset SCZ. Likewise, cortical imbalances in excitatory–inhibitory signaling may result from a cell-autonomous defect in cortical interneuron (cIN) development. Overall, these findings indicate that genetic risk in SCZ impacts neocorticogenesis, hippocampal circuit formation, and the development of distinct glial and neuronal subtypes. In light of this remarkable progress, we discuss current limitations and further steps necessary to harvest the full potential of iPSC-based investigations on psychiatric disorders. Psychiatric disorders are among the conditions with the largest disease burden  and rankThe intricacy of the human brain, its inaccessibility in life, and a shortage of neuropathological clues to disease mechanism have limited research on psychiatric disorders from the very beginning. This dilemma has somewhat improved with the advent of structural magnetic resonance imaging in which patients with SCZ revealed thinning of prefrontal and temporal cerebral cortices . Though Notwithstanding these uncertainties, it is well-established that SCZ and BD are highly heritable in twin and family studies . TherefoDespite remarkable progress in genetics, psychiatric disorders continue to challenge our ability to link genetic associations with molecular and cellular mechanisms that could inform on cognitive, behavioral, and emotional impairments. In fact, current efforts to translate genetic associations into insights on disease mechanisms have been severely delayed by the dearth of appropriate human model systems for dissecting the polygenic disease architecture of psychiatric disorders in the context of living human cell-types from the central nervous system.Fortunately, this situation has profoundly changed with the transformative discovery  that priFollowing pioneering work by Brennand et al. , the numOverall, these studies present remarkable progress in the field, though they also indicate the need for further improvements. Therefore, we will discuss in this review recent advancements on iPSC studies in psychiatric disorders that seek to address present challenges, both in terms of experimental design and new techniques. Concluding, we also consider present hurdles to overcome for harvesting the full potential of iPSC-based studies in psychiatric disorders.Our insight into the structural, cellular, and circuit-level deviations that manifest in early brain development in patients with SCZ and BD is at present very limited. Recent progress in three-dimensional (3D) culture models holds great potential for producing brain-like structures (hereafter referred to as ‘brain organoids’ or ’spheroids’) from patient-specific iPSCs. Thereby, different cell types are thought to recapitulate anatomically relevant spatial organization, in which cellular connections might more closely track those observed in vivo than in conventional 2D-systems . In factIn a recent study, Stachowiak et al.  investigOverall, this study on forebrain organoids firstly showed aberrant subcortical neurogenesis in vitro in patients with SCZ: (i) Proliferative NPCs were retained in the IZ and gave rise to atypically placed deep subcortical neurons, (ii) TBR1-positive pioneer neurons stayed behind in the IZ and fell short to immigrate into upper cortical layers, and (iii) calretinin-positive interneurons exhibited random directionality of neurites, suggesting perturbed intercortical connectivity. While these results are consistent with those from previous 2D-models on impaired migration, differentiation, and connectivity of iPSC-derived NPCs and neuronal cells from patients with SCZ ,25,26, tDespite the raising interest in iPSC-based modeling of psychiatric disorders, a suitable system to investigate well-defined human neuronal circuitries has been lacking until recently. In fact, almost all current studies ,22 have To model hippocampal circuitry in psychiatric disorders, Sarkar et al.  recently+ and K+ currents, spontaneous and evoked action potentials, and spontaneous excitatory postsynaptic currents) and developed dense electrically active connections  with gene expression profiles closely matching the CA3 and the broader CA3 subfields (latter comprises both CA2 and CA3) as evidenced by single-cell sequencing. By contrast, genes specific to DG neurons were weakly expressed in six-week-old hippocampal cultures. In support of their mature stage, CA3-like, but not DG-like neurons, were electrically active  that triggers the elimination of redundant synapses, while strengthening the remaining ones. This process, called ‘synaptic pruning’, occurs also alongside the refinement of cognitive functions in late adolescence and early adulthood  when SCZTo fill this gap, Sellgren et al.  set up aTo assess pruning activity in SCZ, Sellgren et al.  moved onIn the latter respect, a previous study  has showIntriguingly, treatment with minocycline, a broad-spectrum tetracycline antibiotic, revealed a dose-dependent inhibition of synaptosom engulfment in vitro at clinically relevant doses suggesting a potential treatment of individuals at risk. Consistent with this possibility, analysis of large medical health records with a focus on individuals between the ages of 10 and 18, and with at least one electronically prescribed tetracycline, indicated that treatment at a minimum of 90 days was associated with a significant reduction in the risk for incipient SCZ .Taken together, Sellgren et al. showed that increased hiMG-mediated synaptosomal phagocytosis in vitro are associated with C4 risk variants  and may When considering implications from iPSC-based modeling of psychiatric disorders, it is important to stay aware that genes do not encode for psychopathology. As noted before, chromosomal regions and genes associated with psychiatric disorders are numerous, with each genetic variant conferring only subtle changes at the molecular and cellular level. Most of these alterations are predicted to occur within neurons ,40, thusA number of imaging studies on psychiatric patients have suggested disrupted synaptic connectivity in SCZ, preferentially in the prefrontal cortex ,43. EmerAgainst this background, Windrem et al.  establisOLIG1, OLIG2, SOX10, and ZNF488) and early-stage regulated proteins from myelination . Moreover, a group of disease-linked ion channels and genes regulating synapse development and function were downregulated in hGPCs from patients with COS. Together, these data indicate that reduced myelination in shiverer brains transplanted with hGPCs from patients with COS results from perturbed oligodendrocyte and astrocyte differentiation.RNA-sequencing of in vitro differentiated hGPCs from patients with COS  showed, Any impairment in the differentiation of astrocytes could contribute likewise to the pathogenesis of SCZ given their critical role in synaptic development and function . To testTaken together, this study suggests that cell-autonomous deficits in glial development could contribute substantially to COS pathology. Beyond impaired myelogenesis upon engraftment of patient GPCs, defects in astrocytic maturation led to substantial cognitive and behavioral impairments in recipient mice relative to those counterparts transplanted with GPCs from healthy controls. Although differences in cell viability between case/control samples upon transplantation remained untested , a salieAs a final remark we note that Liu et al.  recently+ concentrations, reduced neuronal firing thresholds, and consequently accentuate network desynchronization. In this situation, agents promoting glial K+ uptake may present an opportunity toward relieving glial maturation defects in COS/SCZ and further studies are warranted to explore this option in chimeric-hGPC mice.Collectively, these findings suggest that dysregulated BMP signaling restricts astrocytic differentiation in vitro of GPCs from patients with COS and impairs functional maturation owing to REST-mediated downregulation of potassium channel genes. The latter event could lead to enhanced interstitial K+) or somatostatin- (SST+) positive subtype, which originate from the medial ganglionic eminence. In vivo, altered GABAergic signaling seems to disrupt cortical γ-oscillations in patients with SCZ and thus contribute to cognitive impairments [Imbalances in excitatory–inhibitory signaling are widely viewed as a mechanism for cortical dysfunction in SCZ owing to an enhanced excitatory glutamatergic and/or a diminished inhibitory GABAergic tone . In support of this hypothesis, integration of postmortem brain transcriptomics of major psychiatric disorders  with sinairments . CognitiTo test this hypothesis, Shao et al.  generateElectrophysiological and optogenetic studies were conducted in vivo and in vitro at seven weeks post transplantation. In a first set of experiments, Shao et al. showed that cINs derived from healthy controls and patients with SCZ developed into cortical neurons closely recapitulating the features of endogenous interneurons, and that both groups expressed functional postsynaptic structures responding to host glutamatergic neurons. Similarly, both groups expressed presynaptic structures for the release of GABA and the inhibition of host cortical neurons. In essence, cINs from both controls and cases undistinguishably developed into authentic and fully functional interneurons in recipient mice.PCDHA2, PCDHA3, PCDHA6, and PCDHA8), where downregulation was specific to cINs from cases, but undetectable in glutamatergic-fated neurons derived from iPSCs of either group.RNA-seq analysis of in vitro differentiated cINs from controls and cases  showed gPCDHA downregulation on early neurodevelopment in vivo, Shao et al. [Pcdha-knockout mice that lacked all Pcdha family members. Histological analysis of prefrontal cINs showed reduced formation and arborization of inhibitory synapses, whereas excitatory synapses were unaffected. Consistent with this finding, a previous report [Pcdha-knockout mice exhibited additional deficits in prepulse inhibition. Since all of these behavioral alterations are thought to mimic clinical phenotypes in SCZ, they prompt the question whether Pcdha-knockout mice matched cINs phenotypes from patients with SCZ in vitro and in vivo, and in postmortem human brain. In fact, somatic neurite, and total branch numbers, as well as total neurite lengths were significantly reduced in vitro in iPSC-derived cINs from patients with SCZ relative to controls. Gain and loss of function experiments further revealed that the deficit in cINs from patients with SCZ could be rescued by treatment in vitro with a PKC inhibitor, or conversely, be induced by knockdown of protocadherin expression in cINs from controls. While cINs fate decision and excitatory synapse formation in vivo did not differ between control- and case-engrafted mice, cINs from cases showed a significant deficit in inhibitory synapse formation relative to controls, thus strengthening the results from in vitro culture. Taken together, these findings indicate that iPSC-derived cINs from patients with SCZ form less inhibitory synapses in the absence of other potential disease-associated alterations in cortical circuitry, an outcome pointing to a cell-autonomous defect. Interestingly, these early developmental cINs phenotypes were recapitulated in a SCZ/control study on postmortem brains: In distinction to early developmental stages in vitro and in vivo, postmortem cINs from patients additionally showed a significant reduction in excitatory synapse formation. Therefore, conditions in vitro would be either not permissive to downregulation of excitatory synapses or this event could reflect a secondary cause in vivo. In light of the small-sized (n = 8 for each condition) postmortem brain collection [To assess the impact of o et al.  turned ts report  had detellection , furtherPCDHA genes. In a complementary approach, Pcdha-knockout mice represented cognitive and behavioral deficits mimicking clinical phenotypes from patients with SCZ. While different lines of evidence from this study converge on an early cell-autonomous defect in cINs, less insight has been gained on the precise genetic and molecular mechanisms involved. Particularly, the potential contribution of current risk loci in SCZ to PCDHA2 regulation remained unsettled owing insufficient sample size. Also, how changes in PCDHA2 relate to other differentially expressed genes between cINs from controls and patients with SCZ needs to be clarified, though these seemed to be only few. Current progress in genome editing techniques [Overall, Shao et al. shed new light on the excitatory–inhibitory imbalance hypothesis in SCZ. Integrated analysis of iPSC-derived cINs from patients with SCZ demonstrated deficits in inhibitory synapse function in models of early brain development in vitro and in vivo, and in postmortem adult human brain. This deficit manifests in a cell-autonomous fashion in vitro, independent of the circuit environment during early brain development in patients with SCZ. At the molecular level, this phenotype resulted, at least in part, from downregulation of chniques  allows dPCDHA genes. This gene family regulates cell–cell adhesion in development and beyond, and Pcdha-knockout mice exhibited behavioral impairments reminiscent of clinical symptoms in patients with SCZ. In aggregate, these studies indicate that genetic risk in SCZ impacts neocorticogenesis, hippocampal circuit formation, and the development of distinct glial and neuronal subtypes. In light of these remarkable findings, we next discuss potential caveats and further steps to be taken in the field of iPSC-based modeling of psychiatric disorders.Present iPSC-based disease modeling has made significant progress in tracing human brain development and early neurodevelopmental deviations in psychiatric disorders. While studies on SCZ are leading, we expect other fields of psychiatry  to follow in time as the field of iPSC-based disease modeling grows and matures. To date, iPSC studies on patients with SCZ suggest that distinct impairments in NPC proliferation, migration, and differentiation in 2D-culture extend to 3D-brain organoids and disrupt neocorticogenesis in vitro. Likewise, the formation of circuit-like CA3/DG hippocampal structures in vitro is impaired in patients with SCZ, pointing to a possible cause of impaired cognitive and behavioral functions in vivo. Beyond neurons, emerging evidence points to perturbed glia development in patients with SCZ. Elegant chimeric-mice studies support altered oligodendrocyte and astrocyte development in vivo in recipients engrafted with GPCs from patients with COS. These alterations are associated with behavioral and emotional impairments mimicking those of patients. As astrocytes guide neuronal maturation, this study too highlights the significance of cell–cell interactions in the pathogenesis of SCZ. Along this line, cortical imbalances in excitatory–inhibitory signaling may result from a cell-autonomous defect in cINs development that originates, to some extent, from the downregulation of Progress on human iPSC-derived cellular models has stimulated the generation of 3D-brain organoids for investigating early brain development in vitro, and by implication, neurodevelopment-related disorders. All of these iPSC-derived brain organoids require external patterning cues to attain control over specific brain regions and types of cells generated, and importantly, over reproducibility. While brain organoids produce a vast array of cells resembling in vivo counterparts, they exhibit high variability that could confound the analysis of case/control studies . In thisAnother concern about brain organoids is the inadequate oxygen and nutrient supply owing the absence of vascularization, which restricts progressively maturation. To promote vascularization, growth factors can be supplemented to nascent organoids or neural organoids can be transplanted into the brains of adult mice . EngraftAlternatively, a practicable fully in vitro system is needed to recapitulate key features in psychiatric disorders as well as in other brain diseases. In this regard, Park et al.  recentlyIdeally, any in vitro cell culture model would capture those abnormalities in patients that arise from multiple cell types, their intricate interactions, and the physiological activity of both local and long-distance neural networks. Presently, we have still to reach out for this goal: Early stages of NPC development, neurogenesis, and differentiation typically prevail in current in vitro models, while rather few touch on initial stages of neural-network formation. In any case, cellular traits characteristic of more mature stages such as postnatal myelination remain, so far, largely elusive (see below). Notwithstanding these reservations, the recent generation of hippocampal circuit-like structures in vitro by Sarkar et al.  brings uIPSC-derived cells from case/control studies can be transplanted into neonate or adult rodent brains to study their properties in a more ‘naturalistic’ environment that is presumably more permissive than in vitro conditions to disease processes involving migration, connectivity, circuit integration, and advanced morphological and electrophysiological maturation. In addition, this approach can be used to identify deficits in distinct behavioral domains in recipient hosts that are relevant to psychopathology, though it does not allow for recapitulating systems-level functionality underlying behavioral, cognitive, and emotional impairments in patients with psychiatric disorders.While clinical symptoms in major psychiatric disorders suggest disintegration of neural circuits, we would like to point out that neurons do not operate outside the world of other cell types present in the living brain. Healthy brain function reflects the intricate balance of interactions between neurons, astrocytes, microglia, and vascular cells, which become compromised during disease. Along this line, Windrem et al.  recentlyNRXN1 owing its large size. NRXN1 encodes for a presynaptic cell-adhesion molecule, called neurexin-1, and structural variations in this gene strongly increase risk for both COS and SCZ. Pak et al. [NRXN1 mutations and showed that in neural differentiated cells, heterozygous loss of NRXN1 specifically impaired the early phase of synaptic transmission. The concept of the ‘tripartite synapse’ highlights the importance of astrocytes as equal partners with neuronal presynaptic and postsynaptic terminals in synaptic processing of information. In light of impaired astroglial function in COS and its strong association with 2p16.3, it would be interesting to investigate astrocytic function in heterozygous NRXN1 mice [The presence of large chromosomal abnormalities in COS does affect a vast number of genes, making it challenging to establish clear cause–effect relationships in neurodevelopment. As an exception to this rule, the 2p16.3 microdeletion presents non-recurrent copy number variations that associate only with deletions of k et al.  recentlyXN1 mice  as well.Apart from glial cells, neurons can be likewise transplanted into mice brain. In this regard, Shao et al.  recentlyTwo recent studies based on human pluripotent stem cells transplanted either cortical progenitors and neurons derived from 2D-monolayer culture  or 3D-brAll in all, iPSC-based disease-modeling of psychiatric disorders has made significant progress over the past years with further improvements in technique and reduction in costs to come in the near future. Though we are still far away to reproduce the complexity of the human brain in vitro or in transplanted animals, current models can offer unpresented insight into the underpinning molecular and cellular mechanisms and how they give rise to the formation of cells, synapses, and circuits from which behavior eventually emerges. Knowledge gained from these studies will not only catalyze new concepts on the classification and etiopathogenesis of psychiatric disorders, but also on the development of new treatments to benefit affected patients and their families in the long term.


Aedes aegypti mosquitoes, however non-vector-dependent routes are also possible. Mechanisms of maternal-fetal transmission remain unknown, and the trans-placental route has been extensively studied in animal models and in human samples. The aim of this review was to summarize recent studies that helped to elucidate the mechanism of CZS in animal models and observational studies. There are still challenges in the diagnosis and prevention of CZS in humans, due to the large gap that remains in translating ZIKV research to clinical practice. Translational research linking governments, local health workers, scientists and industry is fundamental to improve care for mothers and children.In 2015–2016, in the Americas, and especially in northeast Brazil, a significant number of cases of microcephaly and other congenital brain abnormalities were linked with an outbreak of Zika virus (ZIKV) infection in pregnant women. While maternal symptoms of ZIKV are generally mild and self-limiting, clinical presentation in fetuses and newborns infected is extensive and includes microcephaly, decreased cortical development, atrophy and hypoplasia of the cerebellum and cerebellar vermis, arthrogryposis, and polyhydramnios. The term congenital ZIKV syndrome (CZS) was introduced to describe the range of findings associated with maternal-fetal ZIKV transmission. ZIKV is primarily transmitted by  Zika virus (ZIKV) transitioned from a generally unknown entity to one of the most studied viruses worldwide over a one-and-a-half-year period. In 2015–2016, the virus was responsible for an outbreak in the Americas and especially in Brazil. During this outbreak, ZIKV infections in pregnant women were linked for the first time to severe fetal malformations, with an impressive number of cases of microcephaly and other congenital brain abnormalities. In 2015–2016, Brazil reported more than 200,000 probable ZIKV cases and nearly 2,000 cases of microcephaly . Soon afAedes aegypti mosquitoes and was first identified in humans in Uganda around 1952. Between 1952 and 2015 there were two major outbreaks. The first was in Yap, Micronesia in 2007 and the second was in French Polynesia in 2013 [ZIKV is a flavivirus similar to Dengue, West Nile, yellow fever, and Japanese encephalitis viruses. It is an arbovirus transmitted primarily by  in 2013 ,11. ZIKV in 2013 .ZIKV is a member of the TORCH family, which includes Toxoplasma gondii, rubella virus, cytomegalovirus, and herpes simplex virus. This family of pathogens possesses the relatively unusual ability to transmit from a mother to her developing fetus or newborn . PreciseThe term congenital ZIKV syndrome (CZS) has been adopted recently to describe the range of findings associated with maternal-fetal ZIKV transmission . While pThe rapid spread of the ZIKV epidemic and the devastating congenital defects it produced spurred governments, the healthcare industry, academia, physicians, and patient advocates internationally. Vast amounts of knowledge have been obtained on the viral genome, structure, pathogenesis and mechanisms of maternal-fetal transmission of ZIKV. Studies continue to address clinical features, fetal complications, long-term consequences of the infection in humans, as well as routes of infection and pathophysiology of the disease in animal models ,22. SpecIn this review, we focus on a number of recent studies that have helped elucidate the mechanism of CZS using animal models and observational studies in humans, and we reflect on lessons learned from the epidemic now that it has waned.The placenta is the primary organ responsible for nurturing the fetus during development. In human placentas, fetal-derived anchoring and chorionic villi form connections to maternal structures. Anchoring villi possess specialized cells, extravillous trophoblasts (EVTs), at their distal ends. These extravillous trophoblasts specifically invade the maternal decidua basalis and maternal vasculature. Chorionic villi are branching tree-like structures that are bathed in maternal blood of the intervillous space. At the core of these chorionic villi are blood vessels that connect to fetal circulation. The surface chorionic villi is composed of an inner layer of cytotrophoblast (CTB) cells and an outer layer of syncytiotrophoblast (STB) cells, and it mediates exchange of materials between maternal and fetal circulations ,28. OtheThe mouse placenta can be divided into three layers. The outermost layer is the maternal compartment which includes the decidua basalis. The middle layer, referred to as the junctional zone, facilitates placental attachment to the uterus. It contains spongiotrophoblasts, a type of cytotrophoblast, and trophoblast giant cells, which invade the decidua basalis. The inner layer, the labyrinth zone, is the site of nutrient and gas exchange between maternal and fetal compartments. Here, fetal capillaries, lined by fetal endothelial cells, are separated from maternal sinusoids by a layer of mononuclear trophoblasts and a bilayer of multinucleated syncytiotrophoblasts ,30. AlthTo understand how ZIKV overcomes placental host defenses, mouse models were developed. Initially, efforts to study ZIKV vertical transmission in a mouse model were hindered by ZIKV’s inability to infect wild-type mice . This chIfnar1−/- mice) or WT mice treated with a monoclonal antibody against the interferon α/β receptor. A trans-placental route of infection was strongly suggested in this model by staining of placental tissues which revealed ZIKV particles in placental trophoblasts and adjacent fetal endothelial cells. Moreover, apoptosis of trophoblasts, disruption of fetal capillaries, and increased nucleated fetal erythrocytes were visualized, suggesting impairment of basic placental functions with ZIKV infection [Ifnar1−/- females crossed to Ifnar1± males and noted that ZIKV titers were higher in placentas of Ifnar1−/- offspring, whereas Ifnar1± fetuses were resorbed due to abnormal placental labyrinth development in the presence of type I interferon [Initial studies in mice exploring ZIKV vertical transmission utilized mice lacking the interferon α/β receptor  . Further females . Further acetate . TogetheRecently, mouse models have been developed to tackle the problem of studying ZIKV only in artificially immunocompromised states. One model employed direct intrauterine inoculation of ZIKV in an immunocompetent mouse. This model recapitulated fetal neural disease associated with ZIKV, worse outcomes associated with infection at earlier gestational age, and the presence of ZIKV in trophoblasts and fetal endothelial cells, and further demonstrated activation of the type I interferon signaling pathway with ZIKV infection . Anotherin vivo into the placenta remains an open avenue of investigation. .Additional models were used to investigate ZIKV viral entry into cells of the maternal-fetal interface. The TAM family of receptors  are known sites of entry exploited by flaviviruses. However, mice lacking TAM receptors did not show a decrease in ZIKV maternal infection or adverse fetal effects . TIM1 isInnate immune mechanisms in the context of congenital ZIKV infection have been investigated. Inhibition of toll-like receptors (TLRs) 3 and 8 were recently shown to inhibit the cytokine output of ZIKV-infected trophoblasts . Inflammatg16l1, important for autophagosome formation inhibited placental ZIKV infection and vertical transmission in a mouse model [atg16l1 deficiency exclusively in the placenta was sufficient to limit ZIKV infection and vertical transmission. Loss of ZIKV-induced autophagy in the placenta rescued intrauterine growth restriction and limited viral load in fetal brain. Thus, the autophagy pathway activity has physiological significance for determining the outcomes of congenital ZIKV syndrome [The autophagy pathway functions to catabolize intracellular units to produce energy and structural components. Autophagy is recognized to be a key player in host defense through xenophagy, the lysosomal degradation of intracellular pathogens. Autophagy has also been shown to be important in placental defense against pathogens ,49. Howese model ,51. Furtsyndrome –55. FurtMicrocephaly was the first fetal abnormality linked with congenital infection by ZIKV. Subsequently severe cerebral atrophy, ventriculomegaly, and intracranial calcification were also identified, expanding the spectrum of fetal malformations associated with ZIKV infection.Animal models showed that ZIKV infection during pregnancy results in reduced fetal cerebral cortical surface area. It was hypothesized that neural progenitor cells are specifically targeted by the virus. Following infection, ZIKV has an inhibitory effect on the proliferation of these cells and this initiates the sequence of malformations associated with CZS .in vitro model based on neurospheres and cerebral organoids and showed that these structures, when infected with a Brazilian strain of ZIKV, are smaller than uninfected controls and controls infected with an African strain of ZIKV [Another study involving animal models used ZIKV isolated from a febrile case from Brazil to demonstrate a reduced number of cells and thickness of cortical layers. This study also used an  of ZIKV . Neurosp of ZIKV ,59. In l of ZIKV ,61 showeOver the past two years, many studies have investigated ZIKV structure and function to better understand how the virus overcomes placental defenses. ZIKV, like other flaviviruses, possesses an approximately 11-kb single-stranded, positive-sense RNA genome. Translation of this genome produces a single viral polypeptide, which is cleaved by viral and host proteases into functional proteins. The ZIKV genome encodes three structural proteins , and seven nonstructural proteins  ,63. A nuin vitro, through binding and inhibition of the ZIKV NS2B-NS3 protease [Drug repurposing is an attractive strategy in the face of an epidemic, as an existing drug can be quickly put into practice whereas development of a new drug or vaccine may not be completed before the epidemic reaches the end of its natural course. Several studies screened large libraries of existing drugs for candidates to test against ZIKV. For example, Xu et al. identified a caspase inhibitor, emricasan, and several inhibitors of cyclin-dependent kinases that demonstrated neuroprotection during ZIKV infection . Barrowsprotease .in silico screen using a US Food and Drug Administration (FDA)-approved drug library for candidates that target the NS2B-NS3 protease of ZIKV [Other groups focused their studies on drugs with existing approved uses during pregnancy. Viral and host mechanisms can intersect to facilitate ZIKV vertical transmission and thus offer promising points of therapeutic intervention. An excellent example of this is hydroxycholoroquine . Hydroxychloroquine is an antimalarial and anti-rheumatic drug also known to inhibit autophagy. Importantly, Cao et al., demonstrated that pharmacologic inhibition of autophagy with hydroxychloroquine attenuated ZIKV infection in mouse placentas and ameliorated fetal growth restriction associated with ZIKV . The mec of ZIKV . Using m of ZIKV . As hydr of ZIKV –75, it iDespite the tremendous progress in cell culture and animal model work reviewed here and elsewhere ,76–85, tIt has also been a challenge to standardize the diagnosis of CZS. Examination of placentas from suspected or confirmed cases of ZIKV infection is recommended as part of gold-standard care for women and their newborns . HoweverCorrelating gestational age at infection with CZS phenotype has been challenging. Most detailed cases represented first trimester infections with symptomatic disease, which were associated with significant numbers of abortions, stillbirths and neonatal deaths –88,91–95in vitro work.An important consideration is the worldwide variation in antenatal screening availability and management options for women with fetal congenital abnormalities. Of note, it is illegal or highly restricted to obtain an induced abortion in most Latin American countries, including Brazil . These fAs of the end of 2018, a valiant cross-disciplinary effort has provided us with incredible insights into the mechanism of CZS. The epidemic has now waned, stalling clinical trials for many drugs and new vaccines that have been proposed for preventing ZIKV infection and CZS. This review highlights key knowledge that has been gained regarding the mechanism of CZS but also points to clinical challenges surrounding CZS that have not been adequately addressed. With climate change and increasing awareness of the potential for arboviruses to undergo maternal-fetal transmission ,100, fur


Three survey tools had been formally validated. Most CPs perceived that AMS improved patient care , and reduced inappropriate antibiotic use . CPs collaborated with prescribers for infection control  and for uncertain antibiotic treatment . CPs educated patients  and screened guideline-compliance of antimicrobial prescriptions . Guidelines, training, interactions with prescribers, and reimbursement models were major barriers to CP-led AMS implementation. A limited number of validated survey tools are available to assess AMS perceptions and practices of CPs. AMS survey tools require further development to assess stewardship knowledge, stewardship targets, and implementation by CPs.The scope of antimicrobial stewardship (AMS) surveys on community pharmacists (CPs) is uncertain. This study examines the breadth and quality of AMS survey tools measuring the stewardship knowledge, perceptions and practices (KPP) of CPs and analyse survey outcomes. Following PRISMA-ScR checklist and Arksey and O’Malley’s methodological framework seven medical databases were searched. Two reviewers independently screened the literatures, assessed quality of surveys and KPP outcomes were analysed and described. Ten surveys were identified that assessed CPs’ AMS perceptions ( Antimicrobial resistance (AMR) is a pressing concern of community health . Given tAMS aims to improve patient care and decrease health care costs associated with infections through coordinated interventions on the choice, dose, dose-regimen, side effects, drug interactions and allergies of prescribed antibiotics . To faciResearch has shown that CP-led AMS strategies  are effeThe Global Respiratory Infection Partnership (GRIP) framework has included pharmacy leadership as one of the essential parts in the provision of advice to prescribers and patients . CPs areA 2019 systematic review and meta-analysis demonstrated that GP-pharmacist collaborative AMS strategies are effective in reducing antibiotic prescribing (up to 12%) and improving guideline-adherent prescribing (up to 16%) by GPs . NeverthAMS surveys have been widely used in hospital settings to determine the status of AMS implementation  and chanA scoping review methodology ,22,23 waThis review focused on the below specific objectives. To identify the breadth and scope of existing AMS surveys targeting CPs.To assess the quality of and gaps in AMS survey tools measuring knowledge, perceptions and practices of CPs regarding AMS.To identify and analyse the types and range of outcomes reported in AMS surveys.To identify the evidence gaps and recommend future directions of research in relation to improving AMS in community pharmacy.A systematic search was conducted in seven databases of Medline, Embase, Emcare, Pubmed, CINAHL, Web of Science, and Pharmaceutical Abstracts from their inception to 30 October 2018. Databases were accessed through the Monash University library systems. Employed search strategy was: [ AND (knowledge* OR attitude* OR perception* OR belief* OR practice* OR barrier* OR facilitator*) AND (pharmacist* OR community pharmacist*) AND ]. A snowballing search strategy was also used to identify articles cited in relevant review studies. Furthermore, manual search was performed in google scholar and relevant pharmacy journals. Pharmacy journals searched were: International Journal of Clinical Pharmacy, International Journal of Pharmacy Practice, Research in Social and Administrative Pharmacy, Journal of Clinical Pharmacy and Therapeutics, Journal of Pharmacy Practice and Research, European Journal of Hospital Pharmacy, Pharmaceutical Journal, Journal of American Pharmacist Association. Journal of Antimicrobial Chemotherapy was also searched as this journal covers AMS relevant articles involving health professionals including pharmacists. SKS and SP independently selected studies after the screening of titles and abstracts followed by reviewing full-text using the Covidence  platform1A national or cross-sectional survey that explored AMS at community pharmacy context.2Survey participants were limited to CPs of any age and level of experiences.3Surveys that employed a single or multiple outcome measure related to CPs’ knowledge, perceptions, practices, barriers and facilitators concerning AMS.4Full text articles are available.1Qualitative interviews, editorials, reports, case studies and case series;2Any survey that did not include AMS as a topic;3Study conducted in other than primary care;4Articles not written in English language.Data were extracted using a customized and constructed data extraction form. We modified the data extraction template used for our previous systematic review . Data exSKS and CB assessed the quality of surveys as high-, medium- or low- quality using published criteria  Supplem. A 10-poAn evidence synthesis method  was usedSynthesis of quantitative data was based on grouping similar fixed responses into categories. The percentage median and inter-quartile range (IQR) were determined where two or more responses were available in a fixed response category. Categories were grouped into perceived knowledge of AMS; perceptions of AMS and AMS practices. A socioecological framework  was usedWe identified 1860 articles, of which 68 were full text reviewed after the screening of titles and abstracts. Ten surveys ,36,37,38n = 2) or paper-based (n = 8). Ten surveys included responses for a total of 1530 CPs. The response rate of survey studies varied from 12.4–96.6%. Two studies did not report the response rate. Five studies used validated instruments; three of five studies used the same validated instrument. Five studies developed a self-administered questionnaire based on literature reviews but did not report validation studies of the tool except one. Five studies used Cronbach alpha to assess the internal consistency of the questionnaires. Only one survey used exploratory factor analysis (EFA) to evaluate the internal structure and construct validity. Survey data were analysed using descriptive statistics (n = 10), linear regression model (n = 3), univariable and multivariable regression model (n = 2), exploratory factor analysis (n = 1) and inferential statistics (n = 4).All ten studies were cross-sectional surveys. The surveys were administered either online /self-care treatment for an infection where antibiotics were not needed. Most often CPs dispensed antimicrobials with more than the prescribed duration of antibiotics on the patient’s request in some settings.A lower proportion of CPs took part in AMS campaign or AMS awareness initiatives.Reported barriers and facilitators that influenced CPs to conduct AMS are summarised in Major barriers with which the highest proportion of CPs agreed were: lack of training to undertake AMS, access to patient’s records and laboratory data, non-receptive behaviours of GPs to CPs’ intervention on the choice of antibiotics, availability of AMS supportive dispensing guidelines, systems that support interacting with GPs, time-poor settings, remunerations and reimbursement models. Furthermore, CPs believed that their undefined roles in AMS and patients’ lack of understanding of their role in AMS were important barriers to implement AMS in community pharmacy.Reported facilitators to improve AMS practices by CPs included their familiarity with AMS, willingness to participate AMS programs, positive intention to collaborate with GPs, public awareness campaign, accessibility to antibiotic guidelines, and patients’ clinical and laboratory reports. Most CPs believed that the provision of AMS education, defining their roles in AMS and financial incentives l would facilitate AMS activities in community pharmacy.There exists a small, but growing body of literature concerning CP-AMS. Ten surveys were found from seven countries globally that measured perceptions and practices of CPs in relation to AMS. The dearth of survey studies may be due to suboptimal progress of AMS implementation in the community compared to hospitals settings. Only three validated surveys were found to assess CPs’ perceptions and practices but no validated survey assessed AMS knowledge. Local validation of these survey tools is required to ensure their suitability and validity within local contexts.The existing AMS surveys are suboptimal to precisely monitor and evaluate the progress of AMS in community pharmacy in line with the country specific national AMR action plan. Some stewardship components could be included to optimally design AMS surveys in future. Therefore, there is much room for improvement in the design of surveys and survey questions to investigate the availability of AMS implementation resources, guidelines, interprofessional collaborative structures and staff required to perform AMS by CPs. Questions related to the changes in infrastructures, organizational setup and pharmacy owners’ attitudes and motivations to support AMS programs could also be included. Additionally, CPs’ attitudes towards GP-pharmacist collaboration strategies, and attitudes towards the feasibility of evidence-based AMS strategies in the context where CPs work should be explored to optimally design AMS programs.Though most CPs knew the term of AMS, gaps in their AMS activities were found in this review. CPs had little understanding of what they are supposed to do when dispensing antibiotics, managing infections and providing patient care that is guided by AMS principles. WHO also reported that the roles of CPs are not well defined in policy documents of most countries . SurveyeThis review demonstrated that nearly half  of CPs educated patients about the use of antibiotics and resistance. Exploration is required on how to make sure the availability of patient’s leaflets and guidelines that can be used by CPs to provide patients the self-care advice for infections, antibiotic compliance advice, and the message that inappropriate antibiotics can kill the protective bacteria in the gut, develop diarrhoea and produce secondary infections .Klepser et al. highlighted that a strong GP-CP collaboration is a fundamental strategy to develop sustainable stewardship interventions in the community . The lacTwo studies in Saudi Arabia  and ChinThe longer duration of antibiotic therapy has been reported as a promoter of the development of AMR by two studies ,45. ApprThe context-specific understanding of CPs’ knowledge, perceptions, practices and barriers related to AMS has unique importance to develop and facilitate CP-led AMS interventions. Jamshed et al. highlighted in his review that majority of CPs are still selling antibiotics for unjustified reasons although they have good awareness and knowledge of antibiotic dispensing in low- and middle-income country settings . In thisThis review had some strengths. First, this is the first scoping review which looked at AMS survey of CPs. Second, PRISMA-ScR checklists and a methodological framework ensured the best practice of conducting this review. Third, quality assessment added methodological updates to develop future CP-AMS survey instruments. Fourth, interpretations of quantitative and qualitative data added a bit more understanding of perceptions, practices and barriers to implementing AMS by CPs.This review had some limitations. First, outcomes were based on a limited number of surveys. Second, while interpreting evidence, it faced some methodological limitations: unclear validity of questions posed; variation in sample size and response rate; and non-response bias. The findings may thus overestimate the AMS perceptions and practices of CPs. Third, literature was limited to only English language. Fourth, differences in pharmaceutical legislation and laws of countries where surveys were performed may influence AMS activities of pharmacists.There exists a small but growing body of AMS survey instruments that can be used for exploring CP-AMS. The validity of CP-AMS survey tools at local, national and international contexts requires to be established. There is a room for improving the standard of existing AMS survey tools with evidence-based AMS strategies, stewardship targets, key stewardship components and attitudes towards prescriber-pharmacist collaboration to identify stewardship needs and monitor stewardship progress in community pharmacy. Contextual variations in pharmacy practices prevent us from drawing firm conclusions on the knowledge, perceptions and practices of CPs regarding AMS at this point. With the limited evidence, CPs positively perceived AMS but felt that their AMS practice improvements require AMS training and guidelines, a GP-CP collaborative system structure, and defining their roles to perform AMS. Further surveys could also be designed to gain better qualitative insights into the barriers at individual, practice, system and policy level to conducting AMS by CPs. This review recommends researching the mechanism on how to enhance the engagement of CPs in AMS and develop a GP-CP collaborative AMS implementation model to address AMR in primary care.


This article is the first of four articles designed to explore the complex interrelationship between Autism Spectrum Disorders (ASD); Obsessive compulsive and Related Disorders (OCRD) and Tic Disorders/Tourette's Syndrome (TD/TS). We begin with an overview TD/TS and follow-up with reviews of OCRD and ASD. The final article in this series represents a synthesis of the neurobiological and genetic markers shared by patients presenting with all three syndromes. The goal is to describe the complex endophenotype of these patients in an effort to better define gene markers that underlie these heterogeneous clinical syndromes. Tic disorders (TD) are a collection of hyperkinetic movements that begin in early childhood. Tics are transient for most affected preschool children but a subgroup development persistent movements or progress to develop Tourette Syndrome (TS). TDs as a group display high heritability rates but definitive gene markers still elude us. The difficulty defining genetic markers is in large part due to the diverse neurodevelopmental trajectory, changing topography and typology, development of a broad spectrum of neurocognitive and behavioral complications, and a mixed pattern of psychiatric comorbidities. The focus of this initial paper is on Tic Disorders (TD)/Tourette Syndrome (TS). Our investigation begins with Tic Disorders as a hyperkinetic movement disorder but then progresses to TS as a more complex neuropsychiatric syndrome associated with multiple, comorbid psychiatric and neurodevelopmental disorders. This level of complexity is in large part related to the tendency of both tics and associated clinical symptoms to evolve along a continuum that moves from largely involuntary movements to a more complex mixture of sensorimotor, affective, behavioral, neurocognitive and neuropsychiatric symptoms.The goal of this paper is to present TD/TS in terms of its diverse phenomenology, neurobiology and complex genetics. Subsequent articles in this series will apply this methodology to Autism Spectrum Disorder (ASD) and Obsessive-compulsive and Related Disorders (OCRD). The final article in this series will concentrate on integrating and synthesizing the complex genetics and neurobiology of combinations that includes TD + OCD; TD + ASD; OCD + ASD, and ASD + TD + OCD. The authors selected this approach to better define a set of neurobiological endophenotypes that will enhance our understanding of the core features of these heterogeneous disorders. Although we will not formally discuss Intellectual Disability  in our formulation, we acknowledge the critical role of ID (IDD) in shaping the severity of ASD; contributing to self-injurious behavior (SIB), aggression and stereotypies, and the clinical expression of TD/TS and OCRD 2.Tics are sudden, intrusive, rapid, recurrent, non-rhythmic movements or vocalizations that seem to have no implicit goal or function. Most tics are involuntary and, like other hyperkinetic movement disorders, tend to wax and wane in intensity, worsen during periods of physiological and psychological stress, but improve or disappear during sleep. As a rule, early onset tics begin during the preschool period, especially among children with pre-existing developmental disorders Tic disorders are classified among the Neurodevelopmental Disorders in the DSM-5. The criteria for diagnosing TD requires not only the presence of abnormal movements and vocalizations, but also includes duration and intensity criteria. Symptoms must be sufficient to contribute to functional impairment in social, emotional, and academic or executive skills. Tics should be present by age 18  but in many cases, mild tics may be overlooked. The problem of recognition is compounded by the presence of ID and ASD for reasons that will be explored later. For now, it is sufficient to state that among patients with ASD and ID the age of onset may be less relevant than the age of recognition.Most children who present with tics during their preschool years generally become tic-free within several months. They are rarely diagnosed but would generally fall into the category Other Specified and Unspecified Tic disorders. A smaller percentage of children develop persistent tics that extend beyond 3 months. They are usually diagnosed as either Provisional Tic Disorder, or Tic Disorder, type unspecified 3.There are several important steps in the differential diagnosis of TDs that provide useful clues about the genetic and pathophysiological origins of tics. The first involves differentiating primary from secondary tic disorders. This process centers on the exclusion of abnormal movements associated with clearly defined genetic, metabolic and neurodegenerative movement disorders. The second step involves differentiating tics from other sources of stereotypic movements, tardive and paroxysmal dyskinesias, myoclonus, substance and medication induced and abnormal movements associated with brain trauma 4.TS frequently presents a more complex clinical picture than the other Tic Disorders. These differences are in large part due to greater diversity and more complicated developmental trajectories during childhood and adolescence As stated above, most affected preschool children present with transient tics. Yet embedded in this early onset group are children who will go on to develop chronic motor or vocal tics over the next 2–3 years. A subset of that group will begin with this initial sequence but go on to develop a combination of chronic vocal and motor tics characteristic of TS. Patients with either pattern of tics reach a peak of intensity within about five years of onset. The majority will experience a gradual waning and eventual disappearance of motor and vocal tics by late adolescence or early adulthood. Many of these adults can experience brief but transient resurgences during periods of increased stress Qualitative changes in the complexity of tics and vocalizations also occur among individuals with severe or persistent TS. Complex tics are characterized by repetitive sequences of movements and vocalizations that are usually associated with sensory phenomena (sensations that precede tics) and/or sudden intrusive images and/or urges to complete these repetitive, ritualized, patterns of behaviors. If ignored, these urges and sensations will intensify, terminating only when the “voluntary” actions are completed or “feels right.” This combination of intrinsically motivated, “voluntary” movements and behaviors also resembles complex stereotypies, ritualized behaviors, compulsions without obsessions and addiction-like behaviors. In severe TS, echo-phenomena (repetitious motor behaviors) and aberrant social behaviors  can also cause significant psychosocial distress The population of patients with severe TS represents a very small fraction of those children with TD. At present, we have a limited understanding of how and why these changes occur. Clinically many of these patients are also unresponsive to standard treatments. Many resemble patients with ASD, addictions and OC spectrum or related disorders. In addition, many patients with sensory or premonitory tics, mixed sensorimotor abnormalities, or intrusive experiences blur the boundaries with other psychiatric disorders such as PTSD. Others may be misdiagnosed with bipolar disorder (waxing and waning symptoms); psychoses such as schizophrenia  and catatonia . This misattribution is most commonly associated with co-occurring ID or ASD Other repetitive behaviors include destructive and self-injurious behaviors that also possess addiction-like properties. These patterns of repetitive behaviors suggest an expanded role for the ventral cortico-striato thalamic motivational circuitry involved in other habit-related behaviors 5.Tic disorders vary in topography, typology and severity. These changes occur at three levels. The first involves waxing and waning course in which severity changes over time. The second is more likely associated with Tourette Disorder and involves the changing distribution and subtypes of motor and vocal tics. The initial clinical presentation of simple tics involves motor (eye blinking or grimacing) or vocal tics (sniffing and throat clearing). In TS, the pattern of emerging tics characteristically progresses from motor to vocal tics (or vice versa) within a year or so of onset although many patients deviate from this sequence. In most cases, the typologies of both motor and phonic tics will change over time This group is characterized by greater genetic loading , and is characterized by early onset tics, maternal smoking and co-occurring neurodevelopmental disorders. The transformation from simple tics is analogous to metamorphoses from one disorder into other far more complex neuropsychiatric conditions. Unfortunately, we know little about this process other than the likely role of neuronal antibodies, inflammatory activation, cumulative developmental insults, and epigenetic changes in this transformation. Expanding our knowledge base of these “malignant” forms of TS may require us to address tic disorders in terms of co-occurring neuropsychiatric disorders such as other genetic forms of movement disorder, and subtypes of OCRD and ASD Previously we alluded to the artificial dichotomy between voluntary and involuntary movements. Simple tics are involuntary. Support for their involuntary nature come from neurophysiological examination of the premotor cortex and striatum. These studies demonstrate a lack of premotor potentials  that are associated with planned, voluntary movements. The absence of these automatic action potentials may explain why patents with simple motor tics do not appear goal-directed, serve any apparent communicative needs, and frequently go unnoticed Studies using CNV suggest a lack of readiness or pre-movement potentials preceding simple tics that changes with the emergence of sensory or premonitory tics. As described above, the urges intensify and drive what appear to be complex, voluntary movements. Transcranial magnetic stimulation (TMS) reveals a decrease in the Cortical Silent Period, and the decreased Short Interval Intra-Cortical Inhibition Responses. These changes are consistent with increased “arousal”, increased readiness and a lower threshold for sensorimotor activity. The mechanism seems most consistent with a deficit in baseline inhibition and top down regulation that are most likely related to decreased numbers of parvalbumin containing GABA interneurons that help regulate neuronal activity. Tic suppression represents a compensatory override of this system from the prefrontal cortex but unfortunately does not turn off the rising need to move Complex tics are preceded by sensory or premonitory sensations that induce the urge to tic. This pattern of behavior can be misattributed to drug-induced akathisia; craving-like behaviors seen in drug abuse; compulsive behaviors observed in OC-related disorders (non-anxious forms of OCD without obsessions), and in some treatment-resistant stereotypies/rituals Patients with severe TD can vary widely in terms of social, emotional and psychological distress. In these cases, there is an apparent expansion of aberrant neurophysiological activity. For example, patients with tics and classic obsessive-compulsive disorder (anxiety type) present with a different pattern of TMS response that suggests a wider network The polymorphic nature of complex tics frequently overshadows the recognition of tic disorders. In addition, their presence complicates the differential diagnosis by requiring a detailed investigation into other sources for sensory or prodromal experiences; OCD (with obsessions and anxiety) and repetitive, addiction-like behaviors symptoms; intrinsic sensory gating issues that give rise to intrusive visual or somatic experiences, and a range of imitative, or asocial ritualistic behaviors Co-occurrence of TD/TS with ID or ASD creates additional complications. For example, stereotypies, self-injury, ritualistic behaviors and impulsive, asocial behaviors create boundary issues with ASD and ID. These changes are not associated with specific gene markers but are surely driven by yet unknown gene-environmental/epigenetic forces. Like many convergence points, we can describe these far better than we can explain them 6.One major source of diversity and heterogeneity arises from chronic psychological, sexual or physical trauma during early childhood On the surface, it appears that trauma provides a reasonable explanation for stress-related influences on tic disorders The patterns of aberrant development linked to ASD/ID, OCD, and TD are the result of altered neurogenesis, neuroplasticity, maturation of synaptic and axonal stability, and myelination. The impact of trauma-related events represents second or third hits in the emergence of complex TS as well as co-occurring behavioral and comorbid neuropsychiatric disorders 7.TS is classified as a chronic, non-progressive hyperkinetic movement disorder that preserves characteristic traits like a waxing and waning clinical course, intensification during periods of excitement or stress, and lessening during sleep. The capacity to suppress abnormal movements observed in TD/TS is clear-cut from other hyperkinetic movement disorders Movement disorders are also associated with mood disorders and neurocognitive changes observed in subcortical “dementias.” These neurocognitive changes include decreases in motivation, deficits in illicit or cued memory, dysfunctional sensory motor skills, sensory gating, set shifting and executive deficits related to motor speed, sequential skill learning, attention deficits  and adaptation of ongoing patterns of behavior. Embedded in these neurocognitive changes is a lower threshold for habit formation: a reliance on external cues to initiate behaviors  and deficits in the automatization of practiced motor skills 8.TS is highly heritable (∼60%) The search for specific gene markers in TD/TS began with Mendelian models of inheritance . One variation of this model argued that a single gene was responsible for tics and associated conditions- ADHD, OCD and other disruptive behavioral disorders. Support for this model has waned greatly as a result of unsuccessful attempts to explain TD/TS in terms of specific chromosomal loci or candidate genes  HDC, SLITRK1, and CNTNAP2) found in single families or a few cases NRXN1, NLGN4) The genetic smoking guns remain elusive but there is evidence for a few rare, highly penetrant mutations  are involved, there is little definitive evidence for genomic imprinting as an explanation for gender dimorphism . Other neurodevelopmental disorders display a similar pattern of gender dimorphism 9.TD/TS are complex phenotypes that represent a final common pathway within and between cortico-striato-thalamic-cortical (CSTC) networks. Early on, simple tics are basically involuntary and involve over excitable premotor and motor pathways. The emergence of premonitory or sensory tics broadens the scope of participating neuro-circuits and marks a shift to both involuntary and urge-driven voluntary movements Historically, hyperkinetic movement disorders were lumped into disorders associated with excessive postsynaptic dopamine activity (hyperdopaminergic state). Although dopaminergic dysregulation plays a central role, it is by no means the only neurotransmitter system involved in the neuropharmacology of TD/TS. Abnormal movements result from an imbalance between dopamine/cholinergic/GABAergic activities within CSTC networks. Unfortunately, even this revised model does not fully explain the sensitivity of most tic disorders to low dose neuroleptic drugs, the lower threshold for extrapyramidal symptoms (including akathisia) and the subsequent increased risk for tardive dyskinesia These caveats point to several crucial issues in this paper. As clinicians and researchers, it is important to think of TD/TS as syndromes with many subtypes and phenocopies. The same level of diversity applies to OCD and related disorders. For example, TD shares a genetic relationship with OCRD but is also associated with high rates of comorbidity with ADHD, anxiety and developmental learning disabilities. As a result, it was assumed that ADHD and TD/TS also shared common risk factors and therefore were genetically related syndromes. More recent research challenges this assumption. The evidence suggests that OCRD and TD/TS are interrelated but ADHD is not directly so. The relationship between ADHD and TD/TSD is more consistent with comorbid but independent syndromes except when ADHD is associated with OCRD. This study needs replication but does suggest that the only shared genetic risk between ADHD and TD/TS is related through the OCRD + ADHD connection Lacking a comprehensive explanation for the nearly 70% rates of co-occurrence between TD/TS and ADHD we turn to gender effects- the bias towards males for both disorders. The gender bias is observed in the majority of neurodevelopmental disorders but whether this relationship solves the problem remains to be seen. There is a second caveat. The combination of ADHD and TD/TS has an adverse effect on impulse control, levels of explosive and aggressive behaviors, irritability and emotional outbursts behaviors, and many asocial behaviors usually associated with TD/TS 10.The complexity and integration of these cortico-striato-thalamic-cortical networks creates a “no man's land” for clinicians. Perhaps the biggest challenge involves developing an understanding of the functional neuroanatomy that leads to the typological, typographical and changing severity of tics. This gap widens when the focus expands to include the forces behind symptom changes over time and makes it difficult to predict the developmental trajectory of tic disorders over time. For those with severe TD/TS there is a limited understanding of the basic functional neuroanatomical changes that contribute to the emergence of challenging behaviors , sensorimotor and neurocognitive symptoms and comorbid mental disorders Tic disorders represent a complex genetic disorder with heterogeneous clinical phenotypes. The level of heritability in part varies with age of onset, gender, comorbidity with ADHD and OCRD, and relationship to neurological, immunological, neurodevelopmental and other movement disorders 11.This paper addresses the complex neurobiology of TD/TS and provides a glimpse into a family of comorbid and co-occurring neuropsychiatric disorders. It outlines some of the areas in which descriptive/categorical diagnostic criteria are less helpful and emphasize the increasing need to define and refine neurobiological endophenotypes. Subsequent articles will explore the many relationships between OCRD, ASD and TD/TD. One primary focus is on each syndrome as a complex genetic disorder that share overlapping functional neuroanatomy, neuropharmacology and clinical symptomatology.The greatest barrier to TD/TS research lies in its diversity and heterogeneity. This variability arises from the large number of neurological, genetic and medical phenocopies. These problems reinforce our understanding of TS/TD as one pattern of phenotypic expression that must be considered in the context of more generalized sensorimotor disinhibition, imbalance between excitatory/inhibitory networks and cortical dysregulation. Much of the variability encountered by clinical and researchers requires us to consider TS/TD as the expression of multiple independent but co-occurring disorders; the product of polygenic inheritance whose phenotypic expression varies in relation to the particular mixture of genes and gene-environment interactions, and the effects of gender on phenotypic expression One approach to address these issues was provided by NIMH in 2013- the Research Domain Criteria


The design of an HIV vaccine remains an elusive but top priority. Data from the non-human primate model and the first moderately protective HIV vaccine trial (RV144) point to a role for qualitative changes in humoral immune functions in protection from infection. Here, we review the current understanding of the antibody response throughout HIV infection, the known correlates of protection, and current strategies to manipulate antibodies to put an end to the epidemic.Recent studies point to innate immune-recruiting antibody function in preventing infection as well as controlling viremia following infection. These data have begun to inform next-generation design of HIV vaccines and antibody therapies by uncovering new viral targets and antibody architectures to improve potency and breadth.Emerging data illustrate a role for innate immune recruiting-antibodies in conferring protection against HIV infection as well as promoting viral control and clearance, offering an unprecedented opportunity to modulate and improve antibody function to fight HIV more effectively. Empirical vaccine design has led to the generation of clinically approved vaccines against 26 pathogens, yet similar approaches have failed against HIV . This isThe first large-scale HIV vaccine trials starting in the late 1980s aimed to induce neutralizing antibodies against the HIV envelope (Env) protein . When vaConcomitantly, a viral vector prime, protein boost strategy was underway using a pox virus prime (ALVAC) and a recombinant Env boost, which resulted in a modest vaccine efficacy of 31.2% , 15, 16.In addition to their role in protection, amassing evidence suggests that antibodies may also contribute to natural control of HIV. Specifically, while only a fraction of spontaneous HIV controllers harbor broad T cell immunity , a largeIn this review, we summarize the knowledge related to the evolution of functional humoral immunity in HIV infection and the correlates of both spontaneous control and vaccine-conferred protection from infection. Finally, we explore how dissecting protective profiles can inform the design of improved HIV vaccines and monoclonal antibody therapies.For most diseases, vaccines are designed to induce antibody responses against the most abundant and immunogenic surface antigen(s) . This isNef gene product drives the rapid downregulation of CD4 from the surface of the infected cell [In order to address this question, discussions have emerged concerning which antigenic targets are most relevant for targeting by non-neutralizing killer antibodies. Upon CD4+ T cell infection, expression of the HIV ted cell . This peted cell . The elited cell . As a reted cell . Given tThis important question remains largely unanswered but may be less concerning in the setting of a protective vaccine response. In this case, few cells are successfully infected , and celDespite these unknowns, emerging data continue to uncover details about the early development of functional antibodies that predict antiviral control and disease progression. Additionally, given the field’s new appreciation for the evolution of neutralizing antibodies in a significant proportion of infected individuals, studies investigating the natural progression of protective humoral immune responses over the course of HIV infection in different populations continue to provide critical clues for the development of vaccines able to leverage protective functions of antibodies.The acute window of HIV infection occurs in the first 2–4 weeks following acquisition. During this time, diagnostic tests often fail to detect infection, as antibodies are still developing during this phase. Importantly, the HIV Env gp160 protein is cleaved intracellularly into two gene products: gp41 that forms the transmembrane region, co-receptor binder, and fusion machinery; and gp120, the extracellular envelope region involved in initial CD4 binding. Although gp41 is more recessed on the viral surface, antibody responses to this epitope evolve first  specific for the autologous infecting HIV strain appear, marking the transition to chronic infection . The virWhile breadth of neutralization evolves in a significant proportion of the infected individuals, several years of infection are required to acquire these protective immune responses. Specifically, evolution of breadth of neutralization occurs over the first 1–3 years of infection, with the percentage of individuals’ neutralizing responses against multiple strains and clades increasing from less than 30 to 75% . EpidemiIn addition to these clinical and immunological correlates, studies have clearly demonstrated an intimate interaction between viral mutational events and neutralizing antibody progression , 50. PerSimilarly to neutralizing antibodies, non-neutralizing antibodies have been observed to influence viral escape and mutation , 60. HowOne of the characteristics of chronic HIV infection is the heterogeneity in disease progression rates across HIV-infected populations , 71. SpeEfforts to define the mechanism(s) that may account for viral control identified an enrichment of genome wide-associated single nucleotide polymorphisms (SNPs) within the major histocompatibility complex (MHC)  in contrTo understand the factors that contribute to viral control beyond genetics and viral infectivity, the field has sought to identify additional immune responses that may be uniquely enriched in ECs Fig. . AnalyseDespite robust evidence supporting a role for ADCC in natural antiviral control, functional antibodies have not yet been shown to provide protection from infection when transferred into NHPs before viral challenge. For example, transfer of polyclonal antibodies from ECs had no impact on HIV acquisition in NHPs. However, titers of these transferred antibodies were associated with a trend towards lower levels of viremia in animal plasma . It reman-acetyl glucosamine residues, thought to be key for promoting enhanced binding to FcγR3a found on NK cells. These data provide evidence of two mechanisms by which controllers selectively enhance Fc-effector activity, via both (1) the selection of more functional antibody subclasses and (2) the generation of antibodies with Fc-glycans engendering enhanced affinity for specific Fc-receptors.Observations of unique functional responses in controllers have prompted deeper investigation into the specific mechanisms that controllers selectively evolve to access broader innate immune activity. Beyond the role of different antibody subclasses in driving enhanced antibody effector function, antibody Fc-glycosylation is also a key modulator of Fc-receptor and complement activation , 86. IntWhile ADCC activity trends higher in controllers , 61–68, Conversely, neutralizing antibodies are largely found in equal or lower frequencies in controllers than in the overall HIV-positive population . Large cOverall, clues from natural infection point to a critical enrichment of functional antibodies in patients who spontaneously control HIV. These data also point to unique approaches to co-evolve nAb and non-nAb activities to leverage both ends of the antibody in HIV prevention.Correlates of protection from infection have emerged from active and passive immunization studies in humans and non-human primates (NHPs) . RemarkaActive immunization in both humans and NHPs repeatedly fail to induce broadly neutralizing antibodies . Still, While only one HIV vaccine has shown promise for protection in humans, a variety of immunization regimens have prevented viral acquisition in NHPs , 105–112Along these lines, passive transfer of broadly neutralizing antibodies (bNAbs) also supports the importance of Fc-effector function in protection. Using a mouse model of HIV infection, some, but not all, bNAbs required Fc-effector function to confer protection from infection . MoreoveThus, like correlates of spontaneous control, insights gleaned from active and passive immunization strategies collectively demonstrate the need to harness both ends of the antibody to fully capitalize on the protective nature of the humoral immune response and prevent HIV infections at a global level.The vast knowledge acquired by the HIV field in the last three decades provides exciting opportunities to guide the design of protective vaccines and therapeutics. These include approaches to leverage both ends of the antibody to enhance both the blockade and killing of the virus and virally infected cells. Despite the successes of pre-exposure prophylaxis (PrEP) and anti-retroviral therapy (ART), a globally protective vaccine remains the simplest and most effective approach to end the HIV epidemic. Adherence to a daily regimen is necessary for the effectiveness of both PrEP and ART, whereas the development and deployment of a durable vaccine is likely to reach a larger fraction of the globe. However, there are still many obstacles to overcome due to the complexity of HIV infection, global diversity, and the subsequent highly heterogenous immune response. Nevertheless, defining correlates of spontaneous HIV control and protection from infection has brought us many steps closer to achieving better control of the disease across the globe and reaching the goal of vaccine design that may leverage these natural immune responses to gain control over the virus. While empirical vaccine design approaches have failed in HIV for the past four decades, emerging correlate-inspired vaccines and therapeutics are certain to revolutionize our fight against HIV.


Nanoparticles (NPs) have found a wide range of applications in clinical therapeutic and diagnostic fields. However, currently most NPs are still in the preclinical evaluation phase with few approved for clinical use. Microfluidic systems can simulate dynamic fluid flows, chemical gradients, partitioning of multi-organs as well as local microenvironment controls, offering an efficient and cost-effective opportunity to fast screen NPs in physiologically relevant conditions. Here, in this review, we are focusing on summarizing key microfluidic platforms promising to mimic in vivo situations and test the performance of fabricated nanoparticles. Firstly, we summarize the key evaluation parameters of NPs which can affect their delivery efficacy, followed by highlighting the importance of microfluidic-based NP evaluation. Next, we will summarize main microfluidic systems effective in evaluating NP haemocompatibility, transport, uptake and toxicity, targeted accumulation and general efficacy respectively, and discuss the future directions for NP evaluation in microfluidic systems. The combination of nanoparticles and microfluidic technologies could greatly facilitate the development of drug delivery strategies and provide novel treatments and diagnostic techniques for clinically challenging diseases. With recent advances in nanotechnology, a variety of nanoparticles (NPs) have been fabricated, including liposomes ,2, gold TM, DoxilTM and gold nanoshellsTM . . 161]. C50 value. This model more mimics the actual microenvironment for tumor progression, invasion, and metastasis, and therefore is more suitable for NP evaluation in terms of anticancer therapy development.Vascularized 3D tumor models could be created in microfluidic systems to better evaluate NP efficacy. Agarwal et al. recently developed a macroscale 3D vascularized tumor model and evaluated NP-mediated drug delivery b 162]. . 162]. TMoreover, microfluidics are unique in detecting the efficacy of shear-activated nanotherapeutics for targeted drug release to obstructed blood vessel areas  Figure c. IntactFinally, new microfluidic systems targeting at parallel testing of multiple drugs or cell types towards personalized medicine and treatment are being developed as well. These systems include single-cell-based assays , 2D cellThe organs-on-a-chip model mimicking one or multiple organs is a shining star in recent years and will continue to be one of the major directions for NP evaluation . It can Currently the interpretation and translation of NP testing results with microfluidic devices to preclinical and clinical outcomes is still a challenge. One reason may be due to the inherent difficulties in recapitulating in vivo functions of organs using various scaling cultures in current microfluidic cultures, residing in almost all in vitro cell and tissue cultures. Another reason may come from the cell source. Currently cell lines are often used in microfluidic systems instead of human primary cells which are difficult to get and maintain the functions. To mimic in vivo human organ and tissue functions, primary cells rather than cell lines, human cells rather than animal cells or tissue slices are preferred. To overcome this in vivo–in vitro correlation challenge, more sophisticated microfluidic systems may be required, which comes together with a better biological understanding of the systems, higher fabrication costs and more comprehensive system monitoring and measuring techniques. At the same time, to push for wider industrial use, high throughput fabrication and screening compatibility of the microfluidic systems are two important factors as well. A balance between model sophistication and model throughput need to be reached for final industrial adoption. On the other hand, standardized microfluidic systems and protocols for specific NP testing application should also be established, which could make NP testing more efficient and comparisons among different systems possible.


Neurodegenerative diseases (NDDs), such as Alzheimer’s disease (AD), Parkinson’s disease (PD), Huntington’s disease (HD), amyotrophic lateral sclerosis (ALS), and frontotemporal dementia (FTD), are devastating age-associated brain disorders. Significant efforts have been made to uncover the molecular and cellular pathogenic mechanisms that underlie NDDs. However, our understanding of the neural circuit mechanisms that mediate NDDs and associated symptomatic features have been hindered by technological limitations. Our inability to identify and track individual neurons longitudinally in subcortical brain regions that are preferentially targeted in NDDs has left gaping holes in our knowledge of NDDs. Recent development and advancement of the miniature fluorescence microscope (miniscope) has opened up new avenues for examining spatially and temporally coordinated activity from hundreds of cells in deep brain structures in freely moving rodents. In the present mini-review, we examine the capabilities of current and future miniscope tools and discuss the innovative applications of miniscope imaging techniques that can push the boundaries of our understanding of neural circuit mechanisms of NDDs into new territories. Neurodegenerative diseases (NDDs), such as Alzheimer’s disease (AD), Parkinson’s disease (PD), Huntington’s disease (HD), amyotrophic lateral sclerosis (ALS), and frontotemporal dementia (FTD), are prevalent and incredibly devastating diseases, with an estimated 6 million people affected by NDDs in the United States alone. A common risk factor for most NDDs is advancing age, and with a globally growing elderly population, the number of cases is expected to increase rapidly worldwide in the coming years. Substantial efforts have been undertaken to study NDDs, which have led to fundamental insights into these diseases. Still, the neurobiology that underlies NDDs is not well understood. With limited knowledge, therapeutic interventions have been developed to delay deterioration and provide temporary relief of symptoms, but there remains a tremendous need to improve treatments.Although NDDs uniquely affect specific neuronal subpopulations, the accumulation of distinct protein-based macroscopic deposits that cause neuronal loss is a common hallmark of NDDs. In parallel with progressive neuronal loss, synaptic dysfunction, altered calcium homeostasis, aberrant neural activity, abnormal neuronal plasticity, and impairment in large-scale neural circuits are well-documented features of many NDDs . The intin vivo calcium imaging technology , a recently developed chnology , to provin vivo calcium transients using microscopy would not be possible without calcium indicators  that leads to an imbalance in basal ganglia direct and indirect pathways . Studies agonist . While tSelective vulnerability of MSNs in the basal ganglia is a characteristic of HD . MSNs arNeuromodulatory system dysfunction has been linked to a number of NDDs, but how specific neuromodulatory systems encode behavior in models of NDD has yet to be examined. Selective loss of cholinergic neurons have been observed in postmortem brains of AD patients , and redγ-aminobutyric acid (GABA) plays a central role in regulating neuronal excitability and maintaining balanced network activity, and inhibitory interneuron abnormalities also contribute to a number of NDDs. Inhibitory interneuron dysfunction has been suggested to contribute to network hypersynchrony  that undA variety of glial cells have complex reciprocal interactions with neurons to support their functions and are active participants in neural circuit refinement . For insThe current generation of miniscopes offers many exciting possibilities for the study of NDDs. Innovation of the miniscope has been dramatically accelerated by open-source sharing of miniscope projects , with fuIn the case of NDDs, it would be advantageous to simultaneously image a preferentially affected cell-type population, as those described above, while also imaging surrounding cell-types. This would provide information to understand how dysfunction of a vulnerable cellular population not only affects behavior, but also the other cells in the local microcircuit. Development of GECIs  with speGECIs with distinct wavelengths also provide an opportunity to concurrently image and manipulate neural activity. Miniscope imaging offers correlational insight into neural activity that encodes behavior, but manipulating neural activity concurrently with imaging can offer insight into causal roles of cell populations within microcircuits . This taFluorescence imaging has focused primarily on measuring intracellular calcium transients. However, florescent sensors for dopamine transients , glutamaExtensive evidence suggests that many NDDs may be characterized by alterations in single-cell firing patterns and dyssynchronization between circuit nodes. Electrophysiological techniques currently offer unparalleled temporal resolution, but are unable to record spatiotemporal dynamics of action potential activity in ensembles with single-neuron resolution . FluoresCapable of performing cell-type specific recordings on large-scale populations longitudinally, the miniscope offers opportunities to ask new and exciting questions in NDDs. Studies utilizing miniscope imaging are increasing rapidly across disciplines of neuroscience but are not commonly being used in NDD models. It is our hope that this mini-review, while by no means exhaustive, brings attention to the powerful capabilities of miniscope imaging, and offers a glimpse into potential questions that could be answered with miniscope experiments in studies of NDDs.CTW, CJW, MF, D-TL, and YL contributed to the preparation of the manuscript. CTW, D-TL, and YL wrote and edited 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.