Nociceptor activity induces nonionotropic NMDA receptor signaling to enable spinal reconsolidation and reverse pathological pain.

Chronic, pathological pain is a highly debilitating condition that can arise and be maintained through central sensitization. Central sensitization shares mechanistic and phenotypic parallels with memory formation. In a sensory model of memory reconsolidation, plastic changes underlying pain hypersensitivity can be dynamically regulated and reversed following the reactivation of sensitized sensory pathways. However, the mechanisms by which synaptic reactivation induces destabilization of the spinal "pain engram" are unclear. We identified nonionotropic N-methyl-d-aspartate receptor (NI-NMDAR) signaling as necessary and sufficient for the reactive destabilization of dorsal horn long-term potentiation and the reversal of mechanical sensitization associated with central sensitization. NI-NMDAR signaling engaged directly or through the reactivation of sensitized sensory networks was associated with the degradation of excitatory postsynaptic proteins. Our findings identify NI-NMDAR signaling as a putative synaptic mechanism by which engrams are destabilized in reconsolidation and as a potential means of treating underlying causes of chronic pain.

Sox2 Ablation in the Suprachiasmatic Nucleus Perturbs Anxiety- and Depressive-like Behaviors.

Mood disorders negatively impact the lives of hundreds of millions of individuals worldwide every year, yet the precise molecular mechanisms by which they manifest remain elusive. Circadian dysregulation is one avenue by which mood disorders are thought to arise. SOX2 is a transcription factor that is highly expressed in the murine suprachiasmatic nucleus (SCN), the circadian master clock, and has been recently found to be an important regulator of Per2, a core component of the molecular clock. Genetic ablation of the Sox2 gene in GABAergic neurons selectively impacts SCN neurons, as they are one of very few, if not the only, GABAergic populations that express Sox2. Here, we show that GABAergic-restricted ablation of Sox2 results in anxio-depressive-like phenotypes in mice as observed in the elevated plus maze, forced swim test, tail suspension test, and sucrose preference test. We further observe a reduction in basal and/or forced swim-induced c-Fos expression, a marker of neuronal activation, in the nucleus incertus, arcuate nucleus, and dentate gyrus of Sox2 conditional knockout (cKO) mice. Given the restricted disruption of SOX2 expression in the SCN of Sox2 cKO mice, we propose that their mood-associated phenotypes are the consequence of a dysregulated central clock that is unable to communicate appropriately timed signals to other brain nuclei that regulate affective behaviors.

SOX2 Regulates Neuronal Differentiation of the Suprachiasmatic Nucleus.

In mammals, the hypothalamic suprachiasmatic nucleus (SCN) functions as the central circadian pacemaker, orchestrating behavioral and physiological rhythms in alignment to the environmental light/dark cycle. The neurons that comprise the SCN are anatomically and functionally heterogeneous, but despite their physiological importance, little is known about the pathways that guide their specification and differentiation. Here, we report that the stem/progenitor cell transcription factor, Sex determining region Y-box 2 (Sox2), is required in the embryonic SCN to control the expression of SCN-enriched neuropeptides and transcription factors. Ablation of Sox2 in the developing SCN leads to downregulation of circadian neuropeptides as early as embryonic day (E) 15.5, followed by a decrease in the expression of two transcription factors involved in SCN development, Lhx1 and Six6, in neonates. Thymidine analog-retention assays revealed that Sox2 deficiency contributed to reduced survival of SCN neurons during the postnatal period of cell clearance, but did not affect progenitor cell proliferation or SCN specification. Our results identify SOX2 as an essential transcription factor for the proper differentiation and survival of neurons within the developing SCN.

Limitations of the Avp-IRES2-Cre (JAX #023530) and Vip-IRES-Cre (JAX #010908) Models for Chronobiological Investigations.

The principal circadian pacemaker in mammals, the suprachiasmatic nucleus (SCN), expresses a number of neuropeptides that facilitate intercellular synchrony, helping to generate coherent outputs to peripheral clocks throughout the body. In particular, arginine vasopressin (AVP)- and vasoactive intestinal peptide (VIP)-expressing neurons have been recognized as crucial subpopulations within the SCN and have thus been the focus of many chronobiological studies. Here, we analyze the neuropeptide expression of 2 popular transgenic mouse strains commonly used to direct or restrict Cre-mediated recombination to AVP- and VIP-ergic neurons. The Avp-IRES2-Cre (JAX #023530) and Vip-IRES-Cre (JAX #010908) "driver" mouse strains express the Cre recombinase under the control of the endogenous Avp or Vip gene, respectively, allowing scientists either to ablate their gene of interest or to overexpress a transgene in a cell type-specific manner. Although these are potentially very powerful tools for chronobiologists and other scientists studying AVP- and VIP-ergic neurons, we found that neuropeptide expression in these mice is significantly decreased when an IRES(2)-Cre cassette is inserted downstream of the neuropeptide-encoding gene locus. The impact of IRES(2)-Cre cassette insertion on neuropeptide expression may be a confounding factor in many experimental designs. Our findings suggest that extreme caution must be exercised when using these mouse models to avoid misinterpretation of empirical results.

SOX2-Dependent Transcription in Clock Neurons Promotes the Robustness of the Central Circadian Pacemaker.

Clock neurons within the mammalian suprachiasmatic nuclei (SCN) encode circadian time using interlocked transcription-translation feedback loops (TTFLs) that drive rhythmic gene expression. However, the contributions of other transcription factors outside of the circadian TTFLs to the functionality of the SCN remain obscure. Here, we report that the stem and progenitor cell transcription factor, sex-determining region Y-box 2 (SOX2), is expressed in adult SCN neurons and positively regulates transcription of the core clock gene, Period2. Mice lacking SOX2 selectively in SCN neurons display imprecise, poorly consolidated behavioral rhythms that do not entrain efficiently to environmental light cycles and that are highly susceptible to constant light-induced arrhythmicity. RNA sequencing revealed that Sox2 deficiency alters the SCN transcriptome, reducing the expression of core clock genes and neuropeptide-receptor systems. By defining the transcriptional landscape within SCN neurons, SOX2 enables the generation of robust, entrainable circadian rhythms that accurately reflect environmental time.

Vocal cord dysfunction following esophageal atresia and tracheoesophageal fistula (EA/TEF) repair.

BACKGROUND: The purpose of this study was to determine risk factors and long-term outcomes in patients with esophageal atresia +/-tracheoesophageal fistula (EA/TEF) with vocal cord dysfunction (VCD) owing to recurrent laryngeal nerve (RLN) injury. METHOD: A retrospective chart review was performed for EA/TEF patients repaired at our institution from 1999 to 2014 (REB #1000032265). RESULTS: Of 197 patients, 22 (11.2%) were diagnosed with VCD by indirect laryngoscopy following EA/TEF repair. Aspiration was documented on video swallow study for 21 patients, and as a result, 13 required thickened feeds and 8 required gastrostomy tube feeds. Of the 16 H-type TEF patients, 8 (50%) had VCD. Following discharge, 20 (90.9%) patients with VCD eventually tolerated full feeds orally without aspiration but only 8 (36.4%) had documented recovery of vocal cord movement at long-term follow up (mean 452 days). Overall, patients with VCD were more likely to have feeding modifications, increased exposure to radiological studies, and increased frequency of Otolaryngology follow-up compared to EA/TEF patients without VCD. CONCLUSION: EA/TEF patients are at risk for VCD. Clinical improvement did not always correlate with recovery of VC motion. Strategies to minimize RLN damage will improve outcomes and quality of life for EA/TEF patients. LEVEL OF EVIDENCE: Level III.