Sensory neuronal sensitisation occurs through HMGB-1-RAGE and TRPV1 in high-glucose conditions.

Many potential causes for painful diabetic neuropathy have been proposed including actions of cytokines and growth factors. High mobility group protein B1 (HMGB1) is a RAGE (also known as AGER) agonist whose levels are increased in diabetes and that contributes to pain by modulating peripheral inflammatory responses. HMGB1 enhances nociceptive behaviour in naïve animals through an unknown mechanism. We tested the hypothesis that HMGB1 causes pain through direct neuronal activation of RAGE and alteration of nociceptive neuronal responsiveness. HMGB1 and RAGE expression were increased in skin and primary sensory (dorsal root ganglion, DRG) neurons of diabetic rats at times when pain behaviour was enhanced. Agonist-evoked TRPV1-mediated Ca2+ responses increased in cultured DRG neurons from diabetic rats and in neurons from naïve rats exposed to high glucose concentrations. HMGB1-mediated increases in TRPV1-evoked Ca2+ responses in DRG neurons were RAGE- and PKC-dependent, and this was blocked by co-administration of the growth factor splice variant VEGF-A165b. Pain behaviour and the DRG RAGE expression increases were blocked by VEGF-A165b treatment of diabetic rats in vivo Hence, we conclude that HMGB1-RAGE activation sensitises DRG neurons in vitro, and that VEGF-A165b blocks HMGB-1-RAGE DRG activation, which may contribute to its analgesic properties in vivo.

Pharmacology of Modulators of Alternative Splicing.

More than 95% of genes in the human genome are alternatively spliced to form multiple transcripts, often encoding proteins with differing or opposing function. The control of alternative splicing is now being elucidated, and with this comes the opportunity to develop modulators of alternative splicing that can control cellular function. A number of approaches have been taken to develop compounds that can experimentally, and sometimes clinically, affect splicing control, resulting in potential novel therapeutics. Here we develop the concepts that targeting alternative splicing can result in relatively specific pathway inhibitors/activators that result in dampening down of physiologic or pathologic processes, from changes in muscle physiology to altering angiogenesis or pain. The targets and pharmacology of some of the current inhibitors/activators of alternative splicing are demonstrated and future directions discussed.

Lamina-specific population encoding of cutaneous signals in the spinal dorsal horn using multi-electrode arrays.

KEY POINTS: Traditional, widely used in vivo electrophysiological techniques for the investigation of spinal processing of somatosensory information fail to account for the diverse functions of each lamina. To overcome this oversimplification, we have used multi-electrode arrays, in vivo, to simultaneously record neuronal activity across all laminae of the spinal dorsal horn. Multi-electrode arrays are sensitive enough to detect lamina- and region-specific encoding of different subtypes of afferent fibres and to detect short-lived changes in synaptic plasticity as measured by the application of cutaneous electrical stimulation of varying intensity and frequency. Differential encoding of innocuous and noxious thermal and mechanical stimuli were also detected across the laminae with the technique, as were the effects of the application of capsaicin. This new approach to the study of the dorsal spinal cord produces significantly more information per experiment, permitting accelerated research whilst also permitting the effects of pharmacological tools to modulate network responses. ABSTRACT: The dorsal horn (DH) of the spinal cord is a complex laminar structure integrating peripheral signals into the central nervous system. Spinal somatosensory processing is commonly measured electrophysiologically in vivo by recording the activity of individual wide-dynamic-range neurons in the deep DH and extrapolating their behaviour to all cells in every lamina. This fails to account for the specialized processes that occur in each lamina and the considerable heterogeneity in cellular phenotype within and between laminae. Here we overcome this oversimplification by employing linear multi-electrode arrays (MEAs) in the DH of anaesthetized rats to simultaneously measure activity across all laminae. The MEAs, comprising 16 channels, were inserted into the lumbar dorsal horn and peripheral neurons activated electrically via transcutaneous electrodes and ethologically with von Frey hairs (vFHs) or an aluminium heating block. Ascending electrical stimuli showed fibre thresholds with distinct dorsoventral innervation profiles. Wind up was observed across the DH during the C-fibre and post-discharge latencies following 0.5 Hz stimulation. Intrathecal application of morphine (5 ng/50 µl) significantly reduced Aδ- and C-fibre-evoked activity in deep and superficial DH. Light vFHs (≤10 g) predominantly activated intermediate and deep laminae whereas noxious vFHs (26 g) also activated the superficial laminae. Noxious heat (55°C) induced significantly greater activity in the superficial and deep laminae than the innocuous control (30°C). The application of these arrays produced the first description of the processing of innocuous and noxious stimuli throughout the intact DH.

VEGFR2 promotes central endothelial activation and the spread of pain in inflammatory arthritis.

Chronic pain can develop in response to conditions such as inflammatory arthritis. The central mechanisms underlying the development and maintenance of chronic pain in humans are not well elucidated although there is evidence for a role of microglia and astrocytes. However in pre-clinical models of pain, including models of inflammatory arthritis, there is a wealth of evidence indicating roles for pathological glial reactivity within the CNS. In the spinal dorsal horn of rats with painful inflammatory arthritis we found both a significant increase in CD11b+ microglia-like cells and GFAP+ astrocytes associated with blood vessels, and the number of activated blood vessels expressing the adhesion molecule ICAM-1, indicating potential glio-vascular activation. Using pharmacological interventions targeting VEGFR2 in arthritic rats, to inhibit endothelial cell activation, the number of dorsal horn ICAM-1+ blood vessels, CD11b+ microglia and the development of secondary mechanical allodynia, an indicator of central sensitization, were all prevented. Targeting endothelial VEGFR2 by inducible Tie2-specific VEGFR2 knock-out also prevented secondary allodynia in mice and glio-vascular activation in the dorsal horn in response to inflammatory arthritis. Inhibition of VEGFR2 in vitro significantly blocked ICAM-1-dependent monocyte adhesion to brain microvascular endothelial cells, when stimulated with inflammatory mediators TNF-α and VEGF-A165a. Taken together our findings suggest that a novel VEGFR2-mediated spinal cord glio-vascular mechanism may promote peripheral CD11b+ circulating cell transmigration into the CNS parenchyma and contribute to the development of chronic pain in inflammatory arthritis. We hypothesise that preventing this glio-vascular activation and circulating cell translocation into the spinal cord could be a new therapeutic strategy for pain caused by rheumatoid arthritis.

Vascular endothelial growth factor-A165b ameliorates outer-retinal barrier and vascular dysfunction in the diabetic retina.

Diabetic retinopathy (DR) is one of the leading causes of blindness in the developed world. Characteristic features of DR are retinal neurodegeneration, pathological angiogenesis and breakdown of both the inner and outer retinal barriers of the retinal vasculature and retinal pigmented epithelial (RPE)-choroid respectively. Vascular endothelial growth factor (VEGF-A), a key regulator of angiogenesis and permeability, is the target of most pharmacological interventions of DR. VEGF-A can be alternatively spliced at exon 8 to form two families of isoforms, pro- and anti-angiogenic. VEGF-A165a is the most abundant pro-angiogenic isoform, is pro-inflammatory and a potent inducer of permeability. VEGF-A165b is anti-angiogenic, anti-inflammatory, cytoprotective and neuroprotective. In the diabetic eye, pro-angiogenic VEGF-A isoforms are up-regulated such that they overpower VEGF-A165b. We hypothesized that this imbalance may contribute to increased breakdown of the retinal barriers and by redressing this imbalance, the pathological angiogenesis, fluid extravasation and retinal neurodegeneration could be ameliorated. VEGF-A165b prevented VEGF-A165a and hyperglycaemia-induced tight junction (TJ) breakdown and subsequent increase in solute flux in RPE cells. In streptozotocin (STZ)-induced diabetes, there was an increase in Evans Blue extravasation after both 1 and 8 weeks of diabetes, which was reduced upon intravitreal and systemic delivery of recombinant human (rh)VEGF-A165b. Eight-week diabetic rats also showed an increase in retinal vessel density, which was prevented by VEGF-A165b. These results show rhVEGF-A165b reduces DR-associated blood-retina barrier (BRB) dysfunction, angiogenesis and neurodegeneration and may be a suitable therapeutic in treating DR.

The control of alternative splicing by SRSF1 in myelinated afferents contributes to the development of neuropathic pain.

Neuropathic pain results from neuroplasticity in nociceptive neuronal networks. Here we demonstrate that control of alternative pre-mRNA splicing, through the splice factor serine-arginine splice factor 1 (SRSF1), is integral to the processing of nociceptive information in the spinal cord. Neuropathic pain develops following a partial saphenous nerve ligation injury, at which time SRSF1 is activated in damaged myelinated primary afferent neurons, with minimal found in small diameter (IB4 positive) dorsal root ganglia neurons. Serine arginine protein kinase 1 (SRPK1) is the principal route of SRSF1 activation. Spinal SRPK1 inhibition attenuated SRSF1 activity, abolished neuropathic pain behaviors and suppressed central sensitization. SRSF1 was principally expressed in large diameter myelinated (NF200-rich) dorsal root ganglia sensory neurons and their excitatory central terminals (vGLUT1+ve) within the dorsal horn of the lumbar spinal cord. Expression of pro-nociceptive VEGF-Axxxa within the spinal cord was increased after nerve injury, and this was prevented by SRPK1 inhibition. Additionally, expression of anti-nociceptive VEGF-Axxxb isoforms was elevated, and this was associated with reduced neuropathic pain behaviors. Inhibition of VEGF receptor-2 signaling in the spinal cord attenuated behavioral nociceptive responses to mechanical, heat and formalin stimuli, indicating that spinal VEGF receptor-2 activation has potent pro-nociceptive actions. Furthermore, intrathecal VEGF-A165a resulted in mechanical and heat hyperalgesia, whereas the sister inhibitory isoform VEGF-A165b resulted in anti-nociception. These results support a role for myelinated fiber pathways, and alternative pre-mRNA splicing of factors such as VEGF-A in the spinal processing of neuropathic pain. They also indicate that targeting pre-mRNA splicing at the spinal level could lead to a novel target for analgesic development.

Vascular Endothelial Growth Factor-A165b Is Protective and Restores Endothelial Glycocalyx in Diabetic Nephropathy.

Diabetic nephropathy is the leading cause of ESRD in high-income countries and a growing problem across the world. Vascular endothelial growth factor-A (VEGF-A) is thought to be a critical mediator of vascular dysfunction in diabetic nephropathy, yet VEGF-A knockout and overexpression of angiogenic VEGF-A isoforms each worsen diabetic nephropathy. We examined the vasculoprotective effects of the VEGF-A isoform VEGF-A165b in diabetic nephropathy. Renal expression of VEGF-A165b mRNA was upregulated in diabetic individuals with well preserved kidney function, but not in those with progressive disease. Reproducing this VEGF-A165b upregulation in mouse podocytes in vivo prevented functional and histologic abnormalities in diabetic nephropathy. Biweekly systemic injections of recombinant human VEGF-A165b reduced features of diabetic nephropathy when initiated during early or advanced nephropathy in a model of type 1 diabetes and when initiated during early nephropathy in a model of type 2 diabetes. VEGF-A165b normalized glomerular permeability through phosphorylation of VEGF receptor 2 in glomerular endothelial cells, and reversed diabetes-induced damage to the glomerular endothelial glycocalyx. VEGF-A165b also improved the permeability function of isolated diabetic human glomeruli. These results show that VEGF-A165b acts via the endothelium to protect blood vessels and ameliorate diabetic nephropathy.

Vascular endothelial growth factor-A165b prevents diabetic neuropathic pain and sensory neuronal degeneration.

Diabetic peripheral neuropathy affects up to half of diabetic patients. This neuronal damage leads to sensory disturbances, including allodynia and hyperalgesia. Many growth factors have been suggested as useful treatments for prevention of neurodegeneration, including the vascular endothelial growth factor (VEGF) family. VEGF-A is generated as two alternative splice variant families. The most widely studied isoform, VEGF-A165a is both pro-angiogenic and neuroprotective, but pro-nociceptive and increases vascular permeability in animal models. Streptozotocin (STZ)-induced diabetic rats develop both hyperglycaemia and many of the resulting diabetic complications seen in patients, including peripheral neuropathy. In the present study, we show that the anti-angiogenic VEGF-A splice variant, VEGF-A165b, is also a potential therapeutic for diabetic neuropathy. Seven weeks of VEGF-A165b treatment in diabetic rats reversed enhanced pain behaviour in multiple behavioural paradigms and was neuroprotective, reducing hyperglycaemia-induced activated caspase 3 (AC3) levels in sensory neuronal subsets, epidermal sensory nerve fibre loss and aberrant sciatic nerve morphology. Furthermore, VEGF-A165b inhibited a STZ-induced increase in Evans Blue extravasation in dorsal root ganglia (DRG), saphenous nerve and plantar skin of the hind paw. Increased transient receptor potential ankyrin 1 (TRPA1) channel activity is associated with the onset of diabetic neuropathy. VEGF-A165b also prevented hyperglycaemia-enhanced TRPA1 activity in an in vitro sensory neuronal cell line indicating a novel direct neuronal mechanism that could underlie the anti-nociceptive effect observed in vivo. These results demonstrate that in a model of Type I diabetes VEGF-A165b attenuates altered pain behaviour and prevents neuronal stress, possibly through an effect on TRPA1 activity.

The degree of acute descending control of spinal nociception in an area of primary hyperalgesia is dependent on the peripheral domain of afferent input.

Descending controls of spinal nociceptive processing play a critical role in the development of inflammatory hyperalgesia. Acute peripheral nociceptor sensitization drives spinal sensitization and activates spino-supraspinal-spinal loops leading to descending inhibitory and facilitatory controls of spinal neuronal activity that further modify the extent and degree of the pain state. The afferent inputs from hairy and glabrous skin are distinct with respect to both the profile of primary afferent classes and the degree of their peripheral sensitization. It is not known whether these differences in afferent input differentially engage descending control systems to different extents or in different ways. Injection of complete Freund's adjuvant resulted in inflammation and swelling of hairy hind foot skin in rats, a transient thermal hyperalgesia lasting <2 h, and longlasting primary mechanical hyperalgesia (≥7 days). Much longer lasting thermal hyperalgesia was apparent in glabrous skin (1 h to >72 h). In hairy skin, transient hyperalgesia was associated with sensitization of withdrawal reflexes to thermal activation of either A- or C-nociceptors. The transience of the hyperalgesia was attributable to a rapidly engaged descending inhibitory noradrenergic mechanism, which affected withdrawal responses to both A- and C-nociceptor activation and this could be reversed by intrathecal administration of yohimbine (α-2-adrenoceptor antagonist). In glabrous skin, yohimbine had no effect on an equivalent thermal inflammatory hyperalgesia. We conclude that acute inflammation and peripheral nociceptor sensitization in hind foot hairy skin, but not glabrous skin, rapidly activates a descending inhibitory noradrenergic system. This may result from differences in the engagement of descending control systems following sensitization of different primary afferent classes that innervate glabrous and hairy skin.

Periaqueductal grey cyclooxygenase-dependent facilitation of C-nociceptive drive and encoding in dorsal horn neurons in the rat.

The experience of pain is strongly affected by descending control systems originating in the brainstem ventrolateral periaqueductal grey (VL-PAG), which control the spinal processing of nociceptive information. A- and C-fibre nociceptors detect noxious stimulation, and have distinct and independent contributions to both the perception of pain quality (fast and slow pain, respectively) and the development of chronic pain. Evidence suggests a separation in the central processing of information arising from A- vs. C-nociceptors; for example, inhibition of the cyclooxygenase-1 (COX-1)-prostaglandin system within the VL-PAG alters spinal nociceptive reflexes evoked by C-nociceptor input in vivo via descending pathways, leaving A-nociceptor-evoked reflexes largely unaffected. As the spinal neuronal mechanisms underlying these different responses remain unknown, we determined the effect of inhibition of VL-PAG COX-1 on dorsal horn wide dynamic-range neurons evoked by C- vs. A-nociceptor activation. Inhibition of VL-PAG COX-1 in anaesthetised rats increased firing thresholds of lamina IV-V wide dynamic-range dorsal horn neurons in response to both A- and C-nociceptor stimulation. Importantly, wide dynamic-range dorsal horn neurons continued to faithfully encode A-nociceptive information, even after VL-PAG COX-1 inhibition, whereas the encoding of C-nociceptor information by wide dynamic-range spinal neurons was significantly disrupted. Dorsal horn neurons with stronger C-nociceptor input were affected by COX-1 inhibition to a greater extent than those with weak C-fibre input. These data show that the gain and contrast of C-nociceptive information processed in individual wide dynamic-range dorsal horn neurons is modulated by prostanergic descending control mechanisms in the VL-PAG.

VEGF-A165b is an endogenous neuroprotective splice isoform of vascular endothelial growth factor A in vivo and in vitro.

Vascular endothelial growth factor (VEGF) A is generated as two isoform families by alternative RNA splicing, represented by VEGF-A165a and VEGF-A165b. These isoforms have opposing actions on vascular permeability, angiogenesis, and vasodilatation. The proangiogenic VEGF-A165a isoform is neuroprotective in hippocampal, dorsal root ganglia, and retinal neurons, but its propermeability, vasodilatatory, and angiogenic properties limit its therapeutic usefulness. In contrast, a neuroprotective effect of endogenous VEGF-A165b on neurons would be advantageous for neurodegenerative pathologies. Endogenous expression of human and rat VEGF-A165b was detected in hippocampal and cortical neurons. VEGF-A165b formed a significant proportion of total VEGF-A in rat brain. Recombinant human VEGF-A165b exerted neuroprotective effects in response to multiple insults, including glutamatergic excitotoxicity in hippocampal neurons, chemotherapy-induced cytotoxicity of dorsal root ganglion neurons, and retinal ganglion cells (RGCs) in rat retinal ischemia-reperfusion injury in vivo. Neuroprotection was dependent on VEGFR2 and MEK1/2 activation but not on p38 or phosphatidylinositol 3-kinase activation. Recombinant human VEGF-A165b is a neuroprotective agent that effectively protects both peripheral and central neurons in vivo and in vitro through VEGFR2, MEK1/2, and inhibition of caspase-3 induction. VEGF-A165b may be therapeutically useful for pathologies that involve neuronal damage, including hippocampal neurodegeneration, glaucoma diabetic retinopathy, and peripheral neuropathy. The endogenous nature of VEGF-A165b expression suggests that non-isoform-specific inhibition of VEGF-A (for antiangiogenic reasons) may be damaging to retinal and sensory neurons.

Vascular endothelial growth factor isoforms differentially protect neurons against neurotoxic events associated with Alzheimer's disease.

Alzheimer's disease (AD) is the most common cause of dementia, the chronic and progressive deterioration of memory and cognitive abilities. AD can be pathologically characterised by neuritic plaques and neurofibrillary tangles, formed by the aberrant aggregation of ß-amyloid and tau proteins, respectively. We tested the hypothesis that VEGF isoforms VEGF-A165a and VEGF-A165b, produced by differential splice site selection in exon 8, could differentially protect neurons from neurotoxicities induced by ß-amyloid and tau proteins, and that controlling expression of splicing factor kinase activity could have protective effects on AD-related neurotoxicity in vitro. Using oxidative stress, ß-amyloid, and tau hyperphosphorylation models, we investigated the effect of VEGF-A splicing isoforms, previously established to be neurotrophic agents, as well as small molecule kinase inhibitors, which selectively inhibit SRPK1, the major regulator of VEGF splicing. While both VEGF-A165a and VEGF-A165b isoforms were protective against AD-related neurotoxicity, measured by increased metabolic activity and neurite outgrowth, VEGF-A165a was able to enhance neurite outgrowth but VEGF-A165b did not. In contrast, VEGF-A165b was more effective than VEGF-A165a in preventing neurite "dieback" in a tau hyperphosphorylation model. SRPK1 inhibition was found to significantly protect against neurite "dieback" through shifting AS of VEGFA towards the VEGF-A165b isoform. These results indicate that controlling the activities of the two different isoforms could have therapeutic potential in Alzheimer's disease, but their effect may depend on the predominant mechanism of the neurotoxicity-tau or ß-amyloid.

Differential roles of galanin on mechanical and cooling responses at the primary afferent nociceptor.

BACKGROUND: Galanin is expressed in a small percentage of intact small diameter sensory neurons of the dorsal root ganglia and in the afferent terminals of the superficial lamina of the dorsal horn of the spinal cord. The neuropeptide modulates nociception demonstrating dose-dependent pro- and anti-nociceptive actions in the naïve animal. Galanin also plays an important role in chronic pain, with the anti-nociceptive actions enhanced in rodent neuropathic pain models. In this study we compared the role played by galanin and its receptors in mechanical and cold allodynia by identifying individual rat C-fibre nociceptors and characterising their responses to mechanical or acetone stimulation. RESULTS: Mechanically evoked responses in C-fibre nociceptors from naive rats were sensitised after close intra-arterial infusion of galanin or Gal2-11 (a galanin receptor-2/3 agonist) confirming previous data that galanin modulates nociception via activation of GalR2. In contrast, the same dose and route of administration of galanin, but not Gal2-11, inhibited acetone and menthol cooling evoked responses, demonstrating that this inhibitory mechanism is not mediated by activation of GalR2. We then used the partial saphenous nerve ligation injury model of neuropathic pain (PSNI) and the complete Freund's adjuvant model of inflammation in the rat and demonstrated that close intra-arterial infusion of galanin, but not Gal2-11, reduced cooling evoked nociceptor activity and cooling allodynia in both paradigms, whilst galanin and Gal2-11 both decreased mechanical activation thresholds. A previously described transgenic mouse line which inducibly over-expresses galanin (Gal-OE) after nerve injury was then used to investigate whether manipulating the levels of endogenous galanin also modulates cooling evoked nociceptive behaviours after PSNI. Acetone withdrawal behaviours in naive mice showed no differences between Gal-OE and wildtype (WT) mice. 7-days after PSNI Gal-OE mice demonstrated a significant reduction in the duration of acetone-induced nociceptive behaviours compared to WT mice. CONCLUSIONS: These data identify a novel galaninergic mechanism that inhibits cooling evoked neuronal activity and nociceptive behaviours via a putative GalR1 mode of action that would also be consistent with a TRP channel-dependent mechanism.

SRPK1 inhibition in vivo: modulation of VEGF splicing and potential treatment for multiple diseases.

SRPK1 (serine-arginine protein kinase 1) is a protein kinase that specifically phosphorylates proteins containing serine-arginine-rich domains. Its substrates include a family of SR proteins that are key regulators of mRNA AS (alternative splicing). VEGF (vascular endothelial growth factor), a principal angiogenesis factor contains an alternative 3' splice site in the terminal exon that defines a family of isoforms with a different amino acid sequence at the C-terminal end, resulting in anti-angiogenic activity in the context of VEGF165-driven neovascularization. It has been shown recently in our laboratories that SRPK1 regulates the choice of this splice site through phosphorylation of the splicing factor SRSF1 (serine/arginine-rich splicing factor 1). The present review summarizes progress that has been made to understand how SRPK1 inhibition may be used to manipulate the balance of pro- and anti-angiogenic VEGF isoforms in animal models in vivo and therefore control abnormal angiogenesis and other pathophysiological processes in multiple disease states.

When Differential Descending Control of Speed Matters: Descending Modulation of A- versus C-Fiber Evoked Spinal Nociception.

Descending pain modulatory systems (DPMS) that originate within the brain and act to modulate spinal nociceptive transmission are a major determinant of the acute and chronic pain experience. Investigations of these systems in basic scientific research is critical to the development of therapeutic strategies for the relief of pain. Despite our best efforts, something is lost in translation. This article will explore whether this is due in part to a primary focus on sensory modality leading to a failure to differentiate between descending control of A- vs. C-fiber mediated spinal nociception.

Hypoxia-induced carbonic anhydrase mediated dorsal horn neuron activation and induction of neuropathic pain.

ABSTRACT: Neuropathic pain, such as that seen in diabetes mellitus, results in part from central sensitisation in the dorsal horn. However, the mechanisms responsible for such sensitisation remain unclear. There is evidence that disturbances in the integrity of the spinal vascular network can be causative factors in the development of neuropathic pain. Here we show that reduced blood flow and vascularity of the dorsal horn leads to the onset of neuropathic pain. Using rodent models (type 1 diabetes and an inducible endothelial-specific vascular endothelial growth factor receptor 2 knockout mouse) that result in degeneration of the endothelium in the dorsal horn, we show that spinal cord vasculopathy results in nociceptive behavioural hypersensitivity. This also results in increased hypoxia in dorsal horn neurons, depicted by increased expression of hypoxia markers such as hypoxia inducible factor 1α, glucose transporter 3, and carbonic anhydrase 7. Furthermore, inducing hypoxia through intrathecal delivery of dimethyloxalylglycine leads to the activation of dorsal horn neurons as well as mechanical and thermal hypersensitivity. This shows that hypoxic signalling induced by reduced vascularity results in increased hypersensitivity and pain. Inhibition of carbonic anhydrase activity, through intraperitoneal injection of acetazolamide, inhibited hypoxia-induced pain behaviours. This investigation demonstrates that induction of a hypoxic microenvironment in the dorsal horn, as occurs in diabetes, is an integral process by which neurons are activated to initiate neuropathic pain states. This leads to the conjecture that reversing hypoxia by improving spinal cord microvascular blood flow could reverse or prevent neuropathic pain.

Activation of the galanin receptor 2 in the periphery reverses nerve injury-induced allodynia.

BACKGROUND: Galanin is expressed at low levels in the intact sensory neurons of the dorsal root ganglia with a dramatic increase after peripheral nerve injury. The neuropeptide is also expressed in primary afferent terminals in the dorsal horn, spinal inter-neurons and in a number of brain regions known to modulate nociception. Intrathecal administration of galanin modulates sensory responses in a dose-dependent manner with inhibition at high doses. To date it is unclear which of the galanin receptors mediates the anti-nociceptive effects of the neuropeptide and whether their actions are peripherally and/or centrally mediated. In the present study we investigated the effects of direct administration into the receptive field of galanin and the galanin receptor-2/3-agonist Gal2-11 on nociceptive primary afferent mechanical responses in intact rats and mice and in the partial saphenous nerve injury (PSNI) model of neuropathic pain. RESULTS: Exogenous galanin altered the responses of mechano-nociceptive C-fibre afferents in a dose-dependent manner in both naive and nerve injured animals, with low concentrations facilitating and high concentrations markedly inhibiting mechano-nociceptor activity. Further, use of the galanin fragment Gal2-11 confirmed that the effects of galanin were mediated by activation of galanin receptor-2 (GalR2). The inhibitory effects of peripheral GalR2 activation were further supported by our demonstration that after PSNI, mechano-sensitive nociceptors in galanin over-expressing transgenic mice had significantly higher thresholds than in wild type animals, associated with a marked reduction in spontaneous neuronal firing and C-fibre barrage into the spinal cord. CONCLUSIONS: These findings are consistent with the hypothesis that the high level of endogenous galanin in injured primary afferents activates peripheral GalR2, which leads to an increase in C-fibre mechanical activation thresholds and a marked reduction in evoked and ongoing nociceptive responses.

Taste perception and implicit attitude toward sweet related to body mass index and soft drink supplementation.

These studies examined the differences in sweet taste perception and implicit attitude toward sweet between normal-weight and overweight/obese adults; and tested the effects of soft drink consumption on sweet taste, explicit preference and implicit attitude toward sweet in normal-weight subjects. In study 1, normal-weight (n = 22) and overweight/obese (n = 11) adults were assessed for sweet taste intensity and pleasantness. Implicit attitude toward sweet was assessed by implicit association test (IAT). In study 2, normal-weight, lightly active adults (n = 12) underwent one month soft drink supplementation (≈760 ml/day). This increased their daily carbohydrate intake by 2.1 ± 0.2g/kg body weight. Sweet taste perception, explicit preference and implicit attitudes to sweet were assessed. In both studies salty taste was also assessed as a contrasting perception. Overweight/obese subjects perceived sweet and salty tastes as less intense (-23% and -19%, respectively) and reported higher IAT scores for sweet than normal-weight controls (2.1-fold). The supplementation changed sweet intensity/pleasantness ratings and it increased explicit preference (2.3-fold) for sweet in a subgroup of initial sucrose-dislikers. In conclusion, overweight/obese individuals are more implicitly attracted to sweet. One month of soft drink supplementation changed sweet taste perception of normal-weight subjects.

The partial saphenous nerve injury model of pain impairs reward-related learning but not reward sensitivity or motivation.

ABSTRACT: Chronic pain is highly comorbid with affective disorders, including major depressive disorder. A core feature of major depressive disorder is a loss of interest in previously rewarding activities. Major depressive disorder is also associated with negative affective biases where cognitive processes are modulated by the affective state. Previous work from our laboratory has shown that reward-related learning and memory is impaired in rodent models of depression generated through a variety of different manipulations. This study investigated different aspects of reward-related behaviour in a rodent model of chronic pain, the partial saphenous nerve injury (PSNI). Using our reward-learning assay, an impairment in reward learning was observed with no difference in sucrose preference, consistent with a lack of effect on reward sensitivity and similar to the effects seen in depression models. In a successive negative contrast task, chronic pain was not associated with changes in motivation for reward either under normal conditions or when reward was devalued although both sham and PSNI groups exhibited the expected negative contrast effect. In the affective bias test, PSNI rats developed a positive affective bias when treated with gabapentin, an effect not seen in the controls suggesting an association with the antinociceptive effects of the drug inducing a relatively more positive affective state. Together, these data suggest that there are changes in reward-related cognition in this chronic pain model consistent with previous findings in rodent models of depression. The effects seen with gabapentin suggest that pain-associated negative affective state may be remediated by this atypical analgesic.