Molecular neurobiological insight into human nervous tissues is needed to generate next-generation therapeutics for neurological disorders such as chronic pain. We obtained human dorsal root ganglia (hDRG) samples from organ donors and performed RNA-sequencing (RNA-seq) to study the hDRG transcriptional landscape, systematically comparing it with publicly available data from a variety of human and orthologous mouse tissues, including mouse DRG (mDRG). We characterized the hDRG transcriptional profile in terms of tissue-restricted gene coexpression patterns and putative transcriptional regulators, and formulated an information-theoretic framework to quantify DRG enrichment. Relevant gene families and pathways were also analyzed, including transcription factors, G-protein-coupled receptors, and ion channels. Our analyses reveal an hDRG-enriched protein-coding gene set (∼140), some of which have not been described in the context of DRG or pain signaling. Most of these show conserved enrichment in mDRG and were mined for known drug-gene product interactions. Conserved enrichment of the vast majority of transcription factors suggests that the mDRG is a faithful model system for studying hDRG, because of evolutionarily conserved regulatory programs. Comparison of hDRG and tibial nerve transcriptomes suggests trafficking of neuronal mRNA to axons in adult hDRG, and are consistent with studies of axonal transport in rodent sensory neurons. We present our work as an online, searchable repository (https://www.utdallas.edu/bbs/painneurosciencelab/sensoryomics/drgtxome), creating a valuable resource for the community. Our analyses provide insight into DRG biology for guiding development of novel therapeutics and a blueprint for cross-species transcriptomic analyses.

SummaryWe generated RNA sequencing data from human DRG samples and comprehensively compared this transcriptome to other human tissues and a matching panel of mouse tissues. Our analysis uncovered functionally enriched genes in the human and mouse DRG with important implications for understanding sensory biology and pain drug discovery. AbstractMolecular neurobiological insight into human nervous tissues is needed to generate next generation therapeutics for neurological disorders like chronic pain. We obtained human Dorsal Root Ganglia (DRG) samples from organ donors and performed RNA-sequencing (RNA-seq) to study the human DRG (hDRG) transcriptional landscape, systematically comparing it with publicly available data from a variety of human and orthologous mouse tissues, including mouse DRG (mDRG). We characterized the hDRG transcriptional profile in terms of tissue-restricted gene co-expression patterns and putative transcriptional regulators, and formulated an information-theoretic framework to quantify DRG enrichment. Our analyses reveal an hDRG-enriched protein-coding gene set (~140), some of which have not been described in the context of DRG or pain signaling. A majority of these show conserved enrichment in mDRG, and were mined for known drug -gene product interactions. Comparison of hDRG and tibial nerve transcriptomes suggest pervasive mRNA transport of sensory neuronal genes to axons in adult hDRG, with potential implications for mechanistic insight into chronic pain in patients. Relevant gene families and pathways were also analyzed, including transcription factors (TFs), g-protein coupled receptors (GCPRs) and ion channels. We present our work as an online, searchable repository (http://www.utdallas.edu/bbs/painneurosciencelab/DRGtranscriptome), creating a valuable resource for the community. Our analyses provide insight into DRG biology for guiding development of novel therapeutics, and a blueprint for cross-species transcriptomic analyses. peer-reviewed)

Background Migraine is characterized by a collection of neurological symptoms in the absence of injury or damage. However, several common preclinical migraine models require significant damage to the skull to stimulate the dura mater, the likely source of afferent signaling leading to head pain. The goal of this study was to determine whether dural stimulation can be performed in mice using an injection that does not cause injury or damage. Methods Using mice, injections of stimuli were administered to the dura mater through the soft tissue at the intersection between the lambdoidal and sagittal sutures. This technique did not require a permanent cannula nor did it cause damage to the skull or dura. Following injection of noxious stimuli, migraine-like behaviors were measured including cutaneous allodynia and facial grimace. The retrograde tracer fluorogold was applied onto the dura using the same injection technique to label trigeminal ganglion cell bodies, which were then testing in vitro using patch-clamp electrophysiology. Results Dural injection of allyl-isothiocyanate, low pH, interleukin-6, or inflammatory soup but not vehicles, led to cephalic/extracephalic allodynia. Facial grimace responses were also observed with allyl-isothiocyanate, pH 6.0, and interleukin-6. Stimulation with interleukin-6 causes priming to normally subthreshold pH 7.0 stimulation of the dura following resolution of the initial interleukin-6 behavior. Systemic injection of sumatriptan at the time of dural stimulation with inflammatory soup decreased the resulting cutaneous hypersensitivity. Trigeminal ganglion cell bodies retrogradely labeled from the dura had low pH-evoked currents similar to those generated by acid-sensing ion channels. Conclusion Non-invasive dural stimulation in mice can be used as a model of migraine in the absence of injury.

Migraine is one of the most common and most disabling disorders. Between attacks, migraine patients are otherwise normal but are sensitized to non-noxious events known as triggers. The purpose of these studies was to investigate whether a headache-like event causes sensitization, or priming, to subsequent subthreshold events. Interleukin-6 (IL-6) was applied to the rat cranial dura mater which produced cutaneous facial and hindpaw allodynia that lasted 24 hours. At 72-hours, IL-6 treated rats developed allodynia in response to dural stimulation with either a pH 6.8 or pH 7.0 solution and to a systemic nitric oxide (NO) donor, a well-known migraine trigger. Vehicle-treated rats did not respond to either pH stimulus nor to the NO donor, demonstrating that IL-6 exposure primes rats to subthreshold stimuli. Inhibitors of brain-derived neurotrophic factor (BDNF) signaling given either systemically or intracisternally 24-hours after IL-6 eliminated responses to dural pH stimulation at 72 hours. Additionally, intracisternal administration of BDNF without prior dural stimulation produced allodynia and once resolved, animals were primed to dural pH 6.8/pH 7.0 and a systemic NO donor. Finally, hindpaw IL-6 produced paw allodynia but not priming to paw injection of pH 7.0 at 72 hours demonstrating differences in priming depending on location. These data indicate that afferent input from the meninges produces BDNF-dependent priming of the dural nociceptive system. This primed state mimics the interictal period of migraine where attacks can be triggered by normally non-noxious events and suggests that BDNF-dependent plasticity may contribute to migraine.

Migraine is one of the most disabling disorders worldwide but the underlying mechanisms are poorly understood. Stress is consistently reported as a common trigger of migraine attacks. Here, we show that repeated stress in mice causes migraine-like behaviors that are responsive to a migraine therapeutic. Adult female and male mice were exposed to 2 hours of restraint stress for 3 consecutive days, after which they demonstrated facial mechanical hypersensitivity and facial grimace responses that were resolved by 14 days after stress. Hypersensitivity or grimace was not observed in either control animals or those stressed for only 1 day. After return to baseline, the nitric oxide donor sodium nitroprusside (SNP; 0.1 mg/kg) elicited mechanical hypersensitivity in stressed but not in control animals, demonstrating the presence of hyperalgesic priming. This suggests the presence of a migraine-like state, because nitric oxide donors are reliable triggers of attacks in migraine patients but not controls. The stress paradigm also caused priming responses to dural pH 7.0 treatment. The presence of this primed state after stress is not permanent because it was no longer present at 35 days after stress. Finally, mice received either the calcitonin gene-related peptide monoclonal antibody ALD405 (10 mg/kg) 24 hours before SNP or a coinjection of sumatriptan (0.6 mg/kg). ALD405, but not sumatriptan, blocked the facial hypersensitivity due to SNP. This stress paradigm in mice and the subsequent primed state caused by stress allow further preclinical investigation of mechanisms contributing to migraine, particularly those caused by common triggers of attacks.

The biological mechanisms supporting age-dependent changes in learning and memory remain elusive. While a growing body of human literature implicates KIBRA in memory and neurodevelopmental disorders, KIBRA’s molecular function and contribution to maturation of synaptic function and cognition remain poorly understood. Despite being expressed throughout early postnatal development, germline deletion of KIBRA impairs synaptic plasticity selectively in adult rodents. However, it is unclear whether KIBRA facilitates proper brain maturation necessary for adult plasticity or whether it plays a distinct role in plasticity in the adult brain. Here, using an inducible KIBRA knockout mouse, we demonstrate that acutely deleting KIBRA in adult forebrain neurons impairs both spatial memory and long-term potentiation (LTP). The deficits in LTP correlate with an adult-selective decrease in extrasynaptic AMPA receptors under basal conditions. We also identify a novel role for KIBRA in LTP-induced AMPAR upregulation. In contrast, acute deletion of KIBRA in juvenile forebrain neurons did not affect LTP and had minimal effects on basal AMPAR expression. These data suggest that KIBRA serves a unique role in adult hippocampal function through regulation of basal and activity-dependent AMPAR proteostasis that supports synaptic plasticity.Significance StatementSynaptic plasticity supported by trafficking of postsynaptic AMPA receptors is a conserved mechanism underlying learning and memory. The nature and efficacy of learning and memory undergo substantial changes during childhood and adolescent development, but the mechanisms underlying this cognitive maturation remain poorly understood. Here, we demonstrate that the human memory- and neurodevelopmental disorder-associated gene KIBRA facilitates memory and hippocampal synaptic plasticity selectively in the adult hippocampus. Furthermore, we show that selective loss of KIBRA from adult but not juvenile neurons reduces expression of extrasynaptic AMPA receptors and prevents LTP-induced increases in AMPAR expression. Overall, our results suggest that KIBRA participates in cellular and molecular processes that become uniquely necessary for memory and synaptic plasticity in early adulthood.

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