Spinal disinhibition: evidence for a hyperpathia phenotype in painful diabetic neuropathy.

The dominant sensory phenotype in patients with diabetic polyneuropathy and neuropathic pain is a loss of function. This raises questions as to which mechanisms underlie pain generation in the face of potentially reduced afferent input. One potential mechanism is spinal disinhibition, whereby a loss of spinal inhibition leads to increased ascending nociceptive drive due to amplification of, or a failure to suppress, incoming signals from the periphery. We aimed to explore whether a putative biomarker of spinal disinhibition, impaired rate-dependent depression of the Hoffmann reflex, is associated with a mechanistically appropriate and distinct pain phenotype in patients with painful diabetic neuropathy. In this cross-sectional study, 93 patients with diabetic neuropathy underwent testing of Hoffmann reflex rate-dependent depression and detailed clinical and sensory phenotyping, including quantitative sensory testing. Compared to neuropathic patients without pain, patients with painful diabetic neuropathy had impaired Hoffmann reflex rate-dependent depression at 1, 2 and 3 Hz (P ≤ 0.001). Patients with painful diabetic neuropathy exhibited an overall loss of function profile on quantitative sensory testing. However, within the painful diabetic neuropathy group, cluster analysis showed evidence of greater spinal disinhibition associated with greater mechanical pain sensitivity, relative heat hyperalgesia and higher ratings of spontaneous burning pain. These findings support spinal disinhibition as an important centrally mediated pain amplification mechanism in painful diabetic neuropathy. Furthermore, our analysis indicates an association between spinal disinhibition and a distinct phenotype, arguably akin to hyperpathia, with combined loss and relative gain of function leading to increasing nociceptive drive.

Sonogenetic control of mammalian cells using exogenous Transient Receptor Potential A1 channels.

Ultrasound has been used to non-invasively manipulate neuronal functions in humans and other animals. However, this approach is limited as it has been challenging to target specific cells within the brain or body. Here, we identify human Transient Receptor Potential A1 (hsTRPA1) as a candidate that confers ultrasound sensitivity to mammalian cells. Ultrasound-evoked gating of hsTRPA1 specifically requires its N-terminal tip region and cholesterol interactions; and target cells with an intact actin cytoskeleton, revealing elements of the sonogenetic mechanism. Next, we use calcium imaging and electrophysiology to show that hsTRPA1 potentiates ultrasound-evoked responses in primary neurons. Furthermore, unilateral expression of hsTRPA1 in mouse layer V motor cortical neurons leads to c-fos expression and contralateral limb responses in response to ultrasound delivered through an intact skull. Collectively, we demonstrate that hsTRPA1-based sonogenetics can effectively manipulate neurons within the intact mammalian brain, a method that could be used across species.

Pharmacological Modulation of Rate-Dependent Depression of the Spinal H-Reflex Predicts Therapeutic Efficacy against Painful Diabetic Neuropathy.

Impaired rate-dependent depression (RDD) of the spinal H-reflex occurs in diabetic rodents and a sub-set of patients with painful diabetic neuropathy. RDD is unaffected in animal models of painful neuropathy associated with peripheral pain mechanisms and diabetic patients with painless neuropathy, suggesting RDD could serve as a biomarker for individuals in whom spinal disinhibition contributes to painful neuropathy and help identify therapies that target impaired spinal inhibitory function. The spinal pharmacology of RDD was investigated in normal rats and rats after 4 and 8 weeks of streptozotocin-induced diabetes. In normal rats, dependence of RDD on spinal GABAergic inhibitory function encompassed both GABAA and GABAB receptor sub-types. The time-dependent emergence of impaired RDD in diabetic rats was preceded by depletion of potassium-chloride co-transporter 2 (KCC2) protein in the dorsal, but not ventral, spinal cord and by dysfunction of GABAA receptor-mediated inhibition. GABAB receptor-mediated spinal inhibition remained functional and initially compensated for loss of GABAA receptor-mediated inhibition. Administration of the GABAB receptor agonist baclofen restored RDD and alleviated indices of neuropathic pain in diabetic rats, as did spinal delivery of the carbonic anhydrase inhibitor acetazolamide. Pharmacological manipulation of RDD can be used to identify potential therapies that act against neuropathic pain arising from spinal disinhibition.

Neuropathic Pain After Spinal Cord Injury: Challenges and Research Perspectives.

Neuropathic pain is a debilitating consequence of spinal cord injury (SCI) that remains difficult to treat because underlying mechanisms are not yet fully understood. In part, this is due to limitations of evaluating neuropathic pain in animal models in general, and SCI rodents in particular. Though pain in patients is primarily spontaneous, with relatively few patients experiencing evoked pains, animal models of SCI pain have primarily relied upon evoked withdrawals. Greater use of operant tasks for evaluation of the affective dimension of pain in rodents is needed, but these tests have their own limitations such that additional studies of the relationship between evoked withdrawals and operant outcomes are recommended. In preclinical SCI models, enhanced reflex withdrawal or pain responses can arise from pathological changes that occur at any point along the sensory neuraxis. Use of quantitative sensory testing for identification of optimal treatment approach may yield improved identification of treatment options and clinical trial design. Additionally, a better understanding of the differences between mechanisms contributing to at- versus below-level neuropathic pain and neuropathic pain versus spasticity may shed insights into novel treatment options. Finally, the role of patient characteristics such as age and sex in pathogenesis of neuropathic SCI pain remains to be addressed.

Analysis of the behavioral, cellular and molecular characteristics of pain in severe rodent spinal cord injury.

Human SCI is frequently associated with chronic pain that is severe and refractory to medical therapy. Most rodent models used to assess pain outcomes in SCI apply moderate injuries to lower thoracic spinal levels, whereas the majority of human lesions are severe in degree and occur at cervical or upper thoracic levels. To better model and understand mechanisms associated with chronic pain after SCI, we subjected adult rats to T3 severe compression or complete transection lesions, and examined pain-related behaviors for three months. Within one week after injury, rats developed consistent forepaw pain-related behaviors including increased spontaneous lifts, tactile allodynia and cold sensitivity that persisted for three months. Place escape avoidance testing confirmed that withdrawal of the forepaws from a von Frey stimulus represented active pain-related aversion. Spontaneous and evoked pain-related measures were attenuated by gabapentin, further indicating that these behaviors reflect development of pain. Spinal level of injury was relevant: rats with T11 severe SCI did not exhibit forepaw pain-related behaviors. Immunoblotting and immunofluorescence of C6-C8 spinal dorsal horn, reflecting sensory innervation of the forepaw, revealed: 1) expansion of CGRP immunoreactivity in lamina I/II; 2) increased GAP-43 expression; and 3) increased IBA1, GFAP and connexin-43 expression. These findings indicate that aberrant pain fiber sprouting and gliopathy occur after severe SCI. Notably, satellite glial cells (SGCs) in C6-C8 DRGs exhibited increases in GFAP and connexin-43, suggesting ongoing peripheral sensitization. Carbenoxolone, a gap junction inhibitor, and specific peptide inhibitors of connexin-43, ameliorated established tactile allodynia after severe SCI. Collectively, severe T3 SCI successfully models persistent pain states and could constitute a useful model system for examining candidate translational pain therapies after SCI.

Induced pluripotent stem cell-derived neural stem cell therapies for spinal cord injury.

The greatest challenge to successful treatment of spinal cord injury is the limited regenerative capacity of the central nervous system and its inability to replace lost neurons and severed axons following injury. Neural stem cell grafts derived from fetal central nervous system tissue or embryonic stem cells have shown therapeutic promise by differentiation into neurons and glia that have the potential to form functional neuronal relays across injured spinal cord segments. However, implementation of fetal-derived or embryonic stem cell-derived neural stem cell therapies for patients with spinal cord injury raises ethical concerns. Induced pluripotent stem cells can be generated from adult somatic cells and differentiated into neural stem cells suitable for therapeutic use, thereby providing an ethical source of implantable cells that can be made in an autologous fashion to avoid problems of immune rejection. This review discusses the therapeutic potential of human induced pluripotent stem cell-derived neural stem cell transplantation for treatment of spinal cord injury, as well as addressing potential mechanisms, future perspectives and challenges.

Remodelling of spared proprioceptive circuit involving a small number of neurons supports functional recovery.

Studies show that limited functional recovery can be achieved by plasticity and adaptation of the remaining circuitry in partial injuries in the central nervous system, although the new circuits that arise in these contexts have not been clearly identified or characterized. We show here that synaptic contacts from dorsal root ganglions to a small number of dorsal column neurons, a caudal extension of nucleus gracilis, whose connections to the thalamus are spared in a precise cervical level 1 lesion, underwent remodeling over time. These connections support proprioceptive functional recovery in a conditioning lesion paradigm, as silencing or eliminating the remodelled circuit completely abolishes the recovered proprioceptive function of the hindlimb. Furthermore, we show that blocking repulsive Wnt signalling increases axon plasticity and synaptic connections that drive greater functional recovery.

Painful neuropathy: Mechanisms.

Painful neuropathy, like the other complications of diabetes, is a growing healthcare concern. Unfortunately, current treatments are of variable efficacy and do not target underlying pathogenic mechanisms, in part because these mechanisms are not well defined. Rat and mouse models of type 1 diabetes are frequently used to study diabetic neuropathy, with rats in particular being consistently reported to show allodynia and hyperalgesia. Models of type 2 diabetes are being used with increasing frequency, but the current literature on the progression of indices of neuropathic pain is variable and relatively few therapeutics have yet been developed in these models. While evidence for spontaneous pain in rodent models is sparse, measures of evoked mechanical, thermal and chemical pain can provide insight into the pathogenesis of the condition. The stocking and glove distribution of pain tantalizingly suggests that the generator site of neuropathic pain is found within the peripheral nervous system. However, emerging evidence demonstrates that amplification in the spinal cord, via spinal disinhibition and neuroinflammation, and also in the brain, via enhanced thalamic activity or decreased cortical inhibition, likely contribute to the pathogenesis of painful diabetic neuropathy. Several potential therapeutic strategies have emerged from preclinical studies, including prophylactic treatments that intervene against underlying mechanisms of disease, treatments that prevent gains of nociceptive function, treatments that suppress enhancements of nociceptive function, and treatments that impede normal nociceptive mechanisms. Ongoing challenges include unraveling the complexity of underlying pathogenic mechanisms, addressing the potential disconnect between the perceived location of pain and the actual pain generator and amplifier sites, and finding ways to identify which mechanisms operate in specific patients to allow rational and individualized choice of targeted therapies.

Altered rate-dependent depression of the spinal H-reflex as an indicator of spinal disinhibition in models of neuropathic pain.

The unpredictable efficacy of current therapies for neuropathic pain may reflect diverse etiological mechanisms operating between, and within, diseases. As descriptions of pain rarely establish specific mechanisms, a tool that can identify underlying causes of neuropathic pain would be useful in developing patient-specific treatments. Rate-dependent depression (RDD), a measure of the change in amplitude of the Hoffman reflex over consecutive stimulations, is attenuated in diabetic rats that also exhibit impaired spinal γ-aminobutyric acid (GABA)A receptor function, reduced spinal potassium chloride co-transporter (KCC2) expression, and indices of painful neuropathy. To investigate whether loss of RDD is a reliable indicator of the contribution of spinal GABAergic dysfunction to neuropathic pain, we assessed RDD, tactile allodynia, and formalin-evoked hyperalgesia in 3 models: rats treated acutely with brain-derived neurotrophic factor (BDNF), diabetic rats treated with the BDNF-sequestering molecule tyrosine receptor kinase B/Fc (TrkB/Fc), and rats with paclitaxel-induced neuropathy. Delivery of BDNF to the spinal cord of normal rats produced RDD deficits and features of painful neuropathy associated with disrupted GABAA receptor-mediated inhibitory function and reduced dorsal spinal KCC2 expression. Treating diabetic rats with TrkB/Fc restored RDD and alleviated indices of painful neuropathy. In paclitaxel-treated rats, RDD was not impaired and behavioral indices of neuropathic pain were not associated with spinal GABAergic dysfunction or reduced dorsal spinal KCC2 expression. Our data reveal BDNF as part of the mechanism underlying spinal cord disinhibition caused by altered GABAA receptor function in diabetic rats and suggest that RDD deficits may be a useful indicator of neuropathic pain states associated with spinal disinhibition, thereby revealing specific therapeutic targets.

Animal models of diabetes-induced neuropathic pain.

Neuropathy will afflict over half of the approximately 350 million people worldwide who currently suffer from diabetes and around one-third of diabetic patients with neuropathy will suffer from painful symptoms that may be spontaneous or stimulus evoked. Diabetes can be induced in rats or mice by genetic, dietary, or chemical means, and there are a variety of well-characterized models of diabetic neuropathy that replicate either type 1 or type 2 diabetes. Diabetic rodents display aspects of sensorimotor dysfunction such as stimulus-evoked allodynia and hyperalgesia that are widely used to model painful neuropathy. This allows investigation of pathogenic mechanisms and development of potential therapeutic interventions that may alleviate established pain or prevent onset of pain.