Faculty Bibliography

BACKGROUND & AIMS/OBJECTIVE:Abdominal pain is a major symptom of diseases that are associated with microbial dysbiosis, including irritable bowel syndrome and inflammatory bowel disease. Germ-free mice are more prone to abdominal pain than conventionally housed mice, and reconstitution of the microbiota in germ-free mice reduces abdominal pain sensitivity. However, the mechanisms underlying microbial modulation of pain remain elusive. We hypothesized that disruption of the intestinal microbiota modulates the excitability of peripheral nociceptive neurons. METHODS:In vivo and in vitro assays of visceral sensation were performed on mice treated with the nonabsorbable antibiotic vancomycin (50 μg/mL in drinking water) for 7 days and water-treated control mice. Bacterial dysbiosis was verified by 16s rRNA analysis of stool microbial composition. RESULTS:Treatment of mice with vancomycin led to an increased sensitivity to colonic distension in vivo and in vitro and hyperexcitability of dorsal root ganglion (DRG) neurons in vitro, compared with controls. Interestingly, hyperexcitability of DRG neurons was not restricted to those that innervated the gut, suggesting a widespread effect of gut dysbiosis on peripheral pain circuits. Consistent with this, mice treated with vancomycin were more sensitive than control mice to thermal stimuli applied to hind paws. Incubation of DRG neurons from naive mice in serum from vancomycin-treated mice increased DRG neuron excitability, suggesting that microbial dysbiosis alters circulating mediators that influence nociception. The cysteine protease inhibitor E64 (30 nmol/L) and the protease-activated receptor 2 (PAR-2) antagonist GB-83 (10 μmol/L) each blocked the increase in DRG neuron excitability in response to serum from vancomycin-treated mice, as did the knockout of PAR-2 in NaV1.8-expressing neurons. Stool supernatant, but not colonic supernatant, from mice treated with vancomycin increased DRG neuron excitability via cysteine protease activation of PAR-2. CONCLUSIONS:Together, these data suggest that gut microbial dysbiosis alters pain sensitivity and identify cysteine proteases as a potential mediator of this effect.

Axons undergo striking changes in their content and distribution of cell adhesion molecules (CAMs) and ion channels during myelination that underlies the switch from continuous to saltatory conduction. These changes include the removal of a large cohort of uniformly distributed CAMs that mediate initial axon-Schwann cell interactions and their replacement by a subset of CAMs that mediate domain-specific interactions of myelinated fibers. Here, using rodent models, we examine the mechanisms and significance of this removal of axonal CAMs. We show that Schwann cells just prior to myelination locally activate clathrin-mediated endocytosis (CME) in axons, thereby driving clearance of a broad array of axonal CAMs. CAMs engineered to resist endocytosis are persistently expressed along the axon and delay both PNS and CNS myelination. Thus, glia non-autonomously activate CME in axons to downregulate axonal CAMs and presumptively axo-glial adhesion. This promotes the transition from ensheathment to myelination while simultaneously sculpting the formation of axonal domains.

Axonal growth cones mediate axonal guidance and growth regulation. We show that migrating neurons in mice possess a growth cone at the tip of their leading process, similar to that of axons, in terms of the cytoskeletal dynamics and functional responsivity through protein tyrosine phosphatase receptor type sigma (PTPσ). Migrating-neuron growth cones respond to chondroitin sulfate (CS) through PTPσ and collapse, which leads to inhibition of neuronal migration. In the presence of CS, the growth cones can revert to their extended morphology when their leading filopodia interact with heparan sulfate (HS), thus re-enabling neuronal migration. Implantation of an HS-containing biomaterial in the CS-rich injured cortex promotes the extension of the growth cone and improve the migration and regeneration of neurons, thereby enabling functional recovery. Thus, the growth cone of migrating neurons is responsive to extracellular environments and acts as a primary regulator of neuronal migration.

PURPOSE OF REVIEW/OBJECTIVE:Continuous renal replacement therapy (CRRT) is a vital medical intervention used in critically ill patients with acute kidney injury (AKI). One of the key components of adequate clearance with CRRT is the use of anticoagulants to prevent clotting of the extracorporeal circuit. Regional citrate anticoagulation is the most often recommended modality. The term 'citrate toxicity' is used to describe potential adverse effects of accumulation of citrate and subsequent hypocalcemia. However, citrate is itself not inherently toxic. The term and diagnosis of citrate toxicity are questioned in this review. RECENT FINDINGS/RESULTS:Citrate is being increasingly used for regional anticoagulation of the CRRT circuit. Citrate accumulation is infrequent and can cause hypocalcemia and metabolic alkalosis, which are potential adverse effects. Citrate itself, however, is not a toxic molecule. The term 'citrate toxicity' has been used to denote hypocalcemia and metabolic acidosis. However, citrate administration is well known to cause systemic and urinary alkalinization and under certain circumstances, metabolic alkalosis, but is not associated itself with any 'toxic' effects.We review the existing literature and debunk the perceived toxicity of citrate. We delve into the metabolism and clearance of citrate and question current data suggesting metabolic acidosis occurs as the result of citrate accumulation. SUMMARY/CONCLUSIONS:In conclusion, this article calls into question prevailing concerns about 'citrate toxicity'. We emphasize the need for a more nuanced understanding of its safety profile. We recommend discarding the term 'citrate toxicity' in favor of another frequently used, but more meaningful term: 'citrate accumulation'.

Neocortical activity is thought to mediate voluntary control over vocal production, but the underlying neural mechanisms remain unclear. In a highly vocal rodent, the male Alston's singing mouse, we investigate neural dynamics in the orofacial motor cortex (OMC), a structure critical for vocal behavior. We first describe neural activity that is modulated by component notes (~100 ms), probably representing sensory feedback. At longer timescales, however, OMC neurons exhibit diverse and often persistent premotor firing patterns that stretch or compress with song duration (~10 s). Using computational modeling, we demonstrate that such temporal scaling, acting through downstream motor production circuits, can enable vocal flexibility. These results provide a framework for studying hierarchical control circuits, a common design principle across many natural and artificial systems.

Psychosocial and environmental factors, including loss of natural reward, contribute to the risk of drug abuse. Reward loss has been modeled in animals by removal from social or sexual contact, transfer from enriched to impoverished housing, or restriction of food. We previously showed that food restriction increases the unconditioned rewarding effects of abused drugs and the conditioned incentive effects of drug-paired environments. Mechanistic studies provided evidence of decreased basal dopamine (DA) transmission, adaptive upregulation of signaling downstream of D1 DA receptor stimulation, synaptic upscaling and incorporation of calcium-permeable AMPA receptors (CP-AMPARs) in medium spiny neurons (MSNs) of nucleus accumbens (NAc). These findings align with the still evolving 'reward deficiency' hypothesis of drug abuse. The present study tested whether a compound natural reward that is known to increase DA utilization, environmental enrichment, would prevent the persistent expression of cocaine conditioned place preference (CPP) otherwise observed in food restricted rats, along with the mechanistic underpinnings. Because nearly all prior investigations of both food restriction and environmental enrichment effects on cocaine CPP were conducted in male rodents, both sexes were included in the present study. Results indicate that environmental enrichment curtailed the persistence of CPP expression, decreased signaling downstream of the D1R, and decreased the amplitude and frequency of spontaneous excitatory postsynaptic currents (EPSCs) in NAc MSNs of food restricted male, but not female, rats. The failure of environmental enrichment to significantly decrease food restriction-induced synaptic insertion of CP-AMPARs, and how this may accord with previous pharmacological findings that blockade of CP-AMPARs reverses behavioral effects of food restriction is discussed. In addition, it is speculated that estrous cycle-dependent fluctuations in DA release, receptor density and MSN excitability may obscure the effect of increased DA signaling during environmental enrichment, thereby interfering with development of the cellular and behavioral effects that enrichment produced in males.

Individuals with Down syndrome (DS) have a partial or complete trisomy of chromosome 21, resulting in an increased risk for early-onset Alzheimer's disease (AD)-type dementia by early midlife. Despite ongoing clinical trials to treat late-onset AD, individuals with DS are often excluded. Furthermore, timely diagnosis or management is often not available. Of the genetic causes of AD, people with DS represent the largest cohort. Currently, there is a knowledge gap regarding the underlying neurobiological mechanisms of DS-related AD (DS-AD), partly due to limited access to well-characterized brain tissue and biomaterials for research. To address this challenge, we created an international consortium of brain banks focused on collecting and disseminating brain tissue from persons with DS throughout their lifespan, named the Down Syndrome Biobank Consortium (DSBC) consisting of 11 biobanking sites located in Europe, India, and the USA. This perspective describes the DSBC harmonized protocols and tissue dissemination goals.

Astrocytes are a heterogeneous population of central nervous system glial cells that respond to pathological insults and injury by undergoing a transformation called "reactivity." Reactive astrocytes exhibit distinct and context-dependent cellular, molecular, and functional state changes that can either support or disturb tissue homeostasis. We recently identified a reactive astrocyte sub-state defined by interferon-responsive genes like Igtp, Ifit3, Mx1, and others, called interferon-responsive reactive astrocytes (IRRAs). To further this transcriptomic definition of IRRAs, we wanted to define the proteomic changes that occur in this reactive sub-state. We induced IRRAs in immunopanned rodent astrocytes and human iPSC-differentiated astrocytes using TNF, IL1α, C1Q, and IFNβ and characterized their proteomic profile (both cellular and secreted) using unbiased quantitative proteomics. We identified 2335 unique cellular proteins, including IFIT2/3, IFITM3, OASL1/2, MX1/2/3, and STAT1. We also report that rodent and human IRRAs secrete PAI1, a serine protease inhibitor which may influence reactive states and functions of nearby cells. Finally, we evaluated how IRRAs are distinct from neurotoxic reactive astrocytes (NRAs). While NRAs are described by expression of the complement protein C3, it was not upregulated in IRRAs. Instead, we found ~90 proteins unique to IRRAs not identified in NRAs, including OAS1A, IFIT3, and MX1. Interferon signaling in astrocytes is critical for the antiviral immune response and for regulating synaptic plasticity and glutamate transport mechanisms. How IRRAs contribute to these functions is unknown. This study provides the basis for future experiments to define the functional roles of IRRAs in the context of neurodegenerative disorders.