ARBEL: A Machine Learning Tool with Light-Based Image Analysis for Automatic Classification of 3D Pain Behaviors.

A detailed analysis of pain-related behaviors in rodents is essential for exploring both the mechanisms of pain and evaluating analgesic efficacy. With the advancement of pose-estimation tools, automatic single-camera video animal behavior pipelines are growing and integrating rapidly into quantitative behavioral research. However, current existing algorithms do not consider an animal's body-part contact intensity with- and distance from- the surface, a critical nuance for measuring certain pain-related responses like paw withdrawals ('flinching') with high accuracy and interpretability.

Identification of novel neuroprotectants against vincristine-induced neurotoxicity in iPSC-derived neurons.

Chemotherapy-induced peripheral neuropathy (CIPN) is a disabling side effect of cancer chemotherapy that can often limit treatment options for cancer patients or have life-long neurodegenerative consequences that reduce the patient's quality of life. CIPN is caused by the detrimental actions of various chemotherapeutic agents on peripheral axons. Currently, there are no approved preventative measures or treatment options for CIPN, highlighting the need for the discovery of novel therapeutics and improving our understanding of disease mechanisms.

Identification of novel neuroprotectants against vincristine-induced neurotoxicity in iPSC-derived neurons.

Chemotherapy-induced peripheral neuropathy (CIPN) is a disabling side effect of cancer chemotherapy that can often limit treatment options for cancer patients or have life-long neurodegenerative consequences that reduce the patient's quality of life. CIPN is caused by the detrimental actions of various chemotherapeutic agents on peripheral axons. Currently, there are no approved preventative measures or treatment options for CIPN, highlighting the need for the discovery of novel therapeutics and improving our understanding of disease mechanisms.

Automated preclinical detection of mechanical pain hypersensitivity and analgesia.

The lack of sensitive and robust behavioral assessments of pain in preclinical models has been a major limitation for both pain research and the development of novel analgesics. Here, we demonstrate a novel data acquisition and analysis platform that provides automated, quantitative, and objective measures of naturalistic rodent behavior in an observer-independent and unbiased fashion. The technology records freely behaving mice, in the dark, over extended periods for continuous acquisition of 2 parallel video data streams: (1) near-infrared frustrated total internal reflection for detecting the degree, force, and timing of surface contact and (2) simultaneous ongoing video graphing of whole-body pose.

Cannabidiol activates neuronal Kv7 channels.

Cannabidiol (CBD), a chemical found in the plant, is a clinically effective antiepileptic drug whose mechanism of action is unknown. Using a fluorescence-based thallium flux assay, we performed a large-scale screen and found enhancement of flux through heterologously expressed human Kv7.2/7.

Topoisomerase I inhibition and peripheral nerve injury induce DNA breaks and ATF3-associated axon regeneration in sensory neurons.

Although axonal damage induces rapid changes in gene expression in primary sensory neurons, it remains unclear how this process is initiated. The transcription factor ATF3, one of the earliest genes responding to nerve injury, regulates expression of downstream genes that enable axon regeneration. By exploiting ATF3 reporter systems, we identify topoisomerase inhibitors as ATF3 inducers, including camptothecin.

Lack of motor recovery after prolonged denervation of the neuromuscular junction is not due to regenerative failure.

Motor axons in peripheral nerves have the capacity to regenerate after injury. However, full functional motor recovery rarely occurs clinically, and this depends on the nature and location of the injury. Recent preclinical findings suggest that there may be a time after nerve injury where, while regrowth to the muscle successfully occurs, there is nevertheless a failure to re-establish motor function, suggesting a possible critical period for synapse reformation.

Robust Axonal Regeneration Occurs in the Injured CAST/Ei Mouse CNS.

Axon regeneration in the CNS requires reactivating injured neurons' intrinsic growth state and enabling growth in an inhibitory environment. Using an inbred mouse neuronal phenotypic screen, we find that CAST/Ei mouse adult dorsal root ganglion neurons extend axons more on CNS myelin than the other eight strains tested, especially when pre-injured. Injury-primed CAST/Ei neurons also regenerate markedly in the spinal cord and optic nerve more than those from C57BL/6 mice and show greater sprouting following ischemic stroke.

Transcriptional profiling at whole population and single cell levels reveals somatosensory neuron molecular diversity.

The somatosensory nervous system is critical for the organism's ability to respond to mechanical, thermal, and nociceptive stimuli. Somatosensory neurons are functionally and anatomically diverse but their molecular profiles are not well-defined. Here, we used transcriptional profiling to analyze the detailed molecular signatures of dorsal root ganglion (DRG) sensory neurons.

Peripheral nerve injury produces a sustained shift in the balance between glutamate release and uptake in the dorsal horn of the spinal cord.

Peripheral nerve injury provokes heightened excitability of primary sensory afferents including nociceptors, and elicits ectopic activity in lesioned and neighboring intact nerve fibers. The major transmitter released by sensory afferents in the superficial dorsal horn of the spinal cord is glutamate. Glutamate is critically involved in nociceptive signaling and the development of neuropathic pain.

TRPA1 contributes to cold hypersensitivity.

TRPA1 is a nonselective cation channel expressed by nociceptors. Although it is widely accepted that TRPA1 serves as a broad irritancy receptor for a variety of reactive chemicals, its role in cold sensation remains controversial. Here, we demonstrate that mild cooling markedly increases agonist-evoked rat TRPA1 currents.

Multiple chronic pain states are associated with a common amino acid-changing allele in KCNS1.

Not all patients with nerve injury develop neuropathic pain. The extent of nerve damage and age at the time of injury are two of the few risk factors identified to date. In addition, preclinical studies show that neuropathic pain variance is heritable.

Cytoplasmic expression systems triggered by mRNA yield increased gene expression in post-mitotic neurons.

Non-viral vectors are promising vehicles for gene therapy but delivery of plasmid DNA to post-mitotic cells is challenging as nuclear entry is particularly inefficient. We have developed and evaluated a hybrid mRNA/DNA system designed to bypass the nuclear barrier to transfection and facilitate cytoplasmic gene expression. This system, based on co-delivery of mRNA(A64) encoding for T7 RNA polymerase (T7 RNAP) with a T7-driven plasmid, produced between 10- and 2200-fold higher gene expression in primary dorsal root ganglion neuronal (DRGN) cultures isolated from Sprague-Dawley rats compared to a cytomegalovirus (CMV)-driven plasmid, and 30-fold greater expression than the enhanced T7-based autogene plasmid pR011.

Schwann cell-derived factor-induced modulation of the NgR/p75NTR/EGFR axis disinhibits axon growth through CNS myelin in vivo and in vitro.

When associated with the Nogo receptor (NgR), the transmembrane receptor p75NTR signals growth cone collapse. Arrest of CNS axon growth in vivo is mediated by CNS myelin-derived inhibitory ligands through either an unknown pathway after NgR- and Ca2+-dependent activation of the epidermal growth factor receptor (EGFR), and/or sequential Rho-A/ROCK/LIM-kinase/cofilin phosphorylation leading to actin depolymerization. Paradoxically, rat retinal ganglion cell (RGC) axons regenerate through the CNS myelin-rich transected optic nerve after intravitreal sciatic nerve grafting without inhibitory ligand neutralization.

A versatile reducible polycation-based system for efficient delivery of a broad range of nucleic acids.

Synthetic vectors based on reducible polycations consisting of histidine and polylysine residues (HIS RPCs) were evaluated for their ability to deliver nucleic acids. Initial experiments showed that RPC-based vectors with at least 70% histidine content mediated efficient levels of gene transfer without requirement for the endosomolytic agent chloroquine. Significant gene transfer was observed in a range of cell types achieving up to a 5-fold increase in the percentage of transfected cells compared to 25 kDa PEI, a gold standard synthetic vector.

Disinhibition of neurotrophin-induced dorsal root ganglion cell neurite outgrowth on CNS myelin by siRNA-mediated knockdown of NgR, p75NTR and Rho-A.

The presence of multiple axon growth inhibitors may partly explain why central nervous system axons are generally incapable of regenerating after injury. Using RNA interference (RNAi) in dorsal root ganglia neurons (DRGN), we demonstrate siRNA-mediated silencing of components of the inhibitory signalling cascade, including p75NTR, NgR and Rho-A mRNA, of 70%, 100% and 100% of the relevant protein, respectively, while changes in neither protein levels nor cellular immunoreactivity were detected using the relevant scrambled siRNA control sequences. Importantly, after 48 h in culture after siRNA-mediated knockdown of Rho-A, neurite outgrowth was enhanced by 30% compared to that after p75NTR and 50% after NgR silencing.

Myelin-, reactive glia-, and scar-derived CNS axon growth inhibitors: expression, receptor signaling, and correlation with axon regeneration.

Axon regeneration is arrested in the injured central nervous system (CNS) by axon growth-inhibitory ligands expressed in oligodendrocytes/myelin, NG2-glia, and reactive astrocytes in the lesion and degenerating tracts, and by fibroblasts in scar tissue. Growth cone receptors (Rc) bind inhibitory ligands, activating a Rho-family GTPase intracellular signaling pathway that disrupts the actin cytoskeleton inducing growth cone collapse/repulsion. The known inhibitory ligands include the chondroitin sulfate proteoglycans (CSPG) Neurocan, Brevican, Phosphacan, Tenascin, and NG2, as either membrane-bound or secreted molecules; Ephrins expressed on astrocyte/fibroblast membranes; the myelin/oligodendrocyte-derived growth inhibitors Nogo, MAG, and OMgp; and membrane-bound semaphorins (Sema) produced by meningeal fibroblasts invading the scar.

Straight talk: new approaches in healthcare. Are you ready to send HIPPA-compliant transactions? Your cash flow is at stake. Panel discussion.

The deadlines are looming for compliance with the transaction and code set requirements set forth in the Health Insurance Portability and Accountability Act of 1996 (HIPAA). If your hospital filed for an extension in October 2002, you need to begin testing transactions by April 2003 and sending transactions by October 2003. But don't rely on your technology vendors to give you the ability to send compliant transactions.

Straight talk: new approaches in health care. HIPAA: deadlines are looming. Are providers prepared?

This is the fourth installment in a series of group discussions by top executives on key issues in healthcare today. Modern Healthcare and PricewaterhouseCoopers present Straight Talk. This session tackles the Health Insurance Portability and Accountability Act of 1996, or HIPAA, and where providers are today in the compliance process and where they need to go.