Model Of Diabetic Neuropathy Research Articles

Blocking mitochondrial calcium release in Schwann cells prevents demyelinating neuropathies.

Schwann cells produce myelin sheath around peripheral nerve axons. Myelination is critical for rapid propagation of action potentials, as illustrated by the large number of acquired and hereditary peripheral neuropathies, such as diabetic neuropathy or Charcot-Marie-Tooth diseases, that are commonly associated with a process of demyelination. However, the early molecular events that trigger the demyelination program in these diseases remain unknown. Here, we used virally delivered fluorescent probes and in vivo time-lapse imaging in a mouse model of demyelination to investigate the underlying mechanisms of the demyelination process. We demonstrated that mitochondrial calcium released by voltage-dependent anion channel 1 (VDAC1) after sciatic nerve injury triggers Schwann cell demyelination via ERK1/2, p38, JNK, and c-JUN activation. In diabetic mice, VDAC1 activity was altered, resulting in a mitochondrial calcium leak in Schwann cell cytoplasm, thereby priming the cell for demyelination. Moreover, reduction of mitochondrial calcium release, either by shRNA-mediated VDAC1 silencing or pharmacological inhibition, prevented demyelination, leading to nerve conduction and neuromuscular performance recovery in rodent models of diabetic neuropathy and Charcot-Marie-Tooth diseases. Therefore, this study identifies mitochondria as the early key factor in the molecular mechanism of peripheral demyelination and opens a potential opportunity for the treatment of demyelinating peripheral neuropathies.

Antinociceptive activity of intraperitoneally administered novel and potent anticonvulsive compound, CY-PROLL-SS, in animal neuropathic pain models

Neuropathic pain is still one of the most difficult pain states to be treated due to the lack of effective drugs. Although the mechanism of action of antiepileptic drugs in alleviating neuropathic pain is not fully understood, it is believed that the bases for both diseases are similar pathophysiologic disturbances. Therefore, in this article we explored the analgesic potential of a recently discovered compound CY-PROLL-SS (ADD 408003; 4-phenyl-perhydropyrrole[1,2-a]pyrazine-1,3-dione) with proved anticonvulsant activity. CY-PROLL-SS was delivered to animals systemically to assess the antinociceptive effects either in streptozocin (STZ)-induced diabetic or in chronic constriction injury (CCI) models of neuropathic pain after acute exposure to both thermal and mechanical stimulus. Examined here compound dose-dependently reversed thermal and mechanical hyperalgesia induced by STZ single injection. Similar results were obtained for CCI-induced hyperalgesia; however, in this case an attenuation of thermal and reversal of mechanical hyperalgesia were observed. High doses of CY-PROLL-SS considerably alleviate peripheral neuropathic pain in model of STZ diabetic neuropathy and CCI. However, mechanisms remain to be elucidated.

Impaired cued and spatial learning performance and altered cannabinoid CB1 receptor functionality in the substantia nigra in a rat model of diabetic neuropathy

Diabetes, and associated diabetic neuropathic pain, impact negatively on cognitive function. However, the underlying mechanisms remain poorly understood. This study investigated neuropathic pain-related behaviour and cognitive function in the rat streptozotocin (STZ) model of diabetes, and assessed cannabinoid1 (CB1) receptor functionality in discrete brain regions. Male Lister-Hooded rats received STZ (60mg/kgs.c.) or vehicle. Sensory responses were assessed in von Frey and Hargreaves tests. Cognitive, motor and sensorimotor functions were assessed using novel object recognition and Morris water maze tasks. CB1 receptor functionality was assessed by [35S]GTPγS (guanosine 5′-O-[gamma-thio]triphosphate) autoradiography. STZ treatment was associated with mechanical allodynia and thermal hypoalgesia. Novel object recognition was unaltered in diabetic rats. STZ treatment was associated with impaired performance in the Morris water maze acquisition phase, but there were no differences in memory retrieval in the probe trial. Stimulus-response learning in the water maze cued trial was also disrupted in STZ-treated rats, possibly indicating sensorimotor deficits. CB1 receptor agonist-stimulated [35S]GTPγS binding was attenuated in the substantia nigra of STZ-treated rats but unaltered in the hippocampus. In conclusion, STZ treatment as a model of diabetic neuropathy was associated with specific functional deficits in the Morris water maze, effects which may be related to altered CB1 receptor functionality in the substantia nigra.

Neuroprotective effects of chronic administration of levetiracetam in a rat model of diabetic neuropathy

ObjectiveDiabetic neuropathy (DNP) is a frequent and serious complication of diabetes mellitus (DM) that leads to progressive and length-dependent loss of peripheral nerve axons. The purpose of the present study is to assess the neuroprotective effects of levetiracetam (LEV) on DNP in a streptozotocin (STZ)-induced DM model in rats. MethodsAdult Sprague-Dawley rats were administered with STZ (60mg/kg) to induce diabetes. DNP was confirmed by electromyography (EMG) and motor function test on 21st day following STZ injection. Study groups were assigned as follows; Group 1: Naïve control (n=8), Group 2: DM+1mL/kg saline (n=12), Group 3: DM+300mg/kg LEV (n=10), Group 4: DM+600mg/kg LEV (n=10). LEV was administered i.p. for 30 consecutive days. Then, EMG, motor function test, biochemical analysis (plasma lipid peroxides and total anti-oxidant capacity), histological and immunohistochemical analysis of sciatic nerves (TUNEL assay, bax, caspase 3, caspase 8 and NGF) were performed to evaluate the efficacy of LEV. ResultsTreatment of diabetic rats with LEV significantly attenuated the inflammation and fibrosis in sciatic nerves and prevented electrophysiological alterations. Immunohistochemistry of sciatic nerves showed a considerable increase in bax, caspase 3 and caspase 8 and a decrease in NGF expression in saline-treated rats whereas LEV significantly suppressed apoptosis markers and prevented the reduction in NGF expression. Besides, LEV considerably reduced plasma lipid peroxides and increased total anti-oxidant capacity in diabetic rats. ConclusionsThe results of the present study suggest that LEV may have therapeutic effects in DNP through modulation of anti-oxidant and anti-apoptotic pathways.

Myelinated 3D neural culture platform as a physiological model of diabetic peripheral neuropathy

Event Abstract Back to Event Myelinated 3D neural culture platform as a physiological model of diabetic peripheral neuropathy Ashwin Sivakumar1, Reed Gioe1 and Michael Moore1, 2 1 Tulane University, Department of Biomedical Engineering, United States 2 Tulane University, Department of Neuroscience, United States Introduction: We present a microengineered 3D hydrogel platform for culture of myelinated embryonic peripheral neural tissue as shown in Figure 1. This novel in-vitro platform enables morphological and physiological metrics analogous to clinical nerve fiber density and nerve conduction tests. We demonstrate the feasibility of this model system for studying peripheral neuropathy caused by glucose-induced neurotoxicity, the most common form of neuropathy. Figure 1: Microengineered myelinated neural fiber tract 3D architecture resembling native peripheral nerve anatomy. Materials and Methods: Dorsal root ganglia (DRG) from embryonic day 15 rats were grown over a period of 4 weeks in a dual hydrogel system consisting of a methacrylated heparin gel in a polyethylene glycol micro-mold. The dual hydrogel system served to constrain the axon growth to be polarized with high density, fasciculation and myelination. The DRG’s were subject to a three-stage media regimen including ascorbic acid (AA) in the final stage to induce Schwann Cell myelination. The myelinated cultures were subjected to 60mM concentrations of the D-glucose in media over a period of 48 hours to simulate diabetic neuropathy. The control cultures were exposed to a 25mM D-glucose concentration. Electrophysiological field recordings were taken with and without the exposure of high glucose, and changes in the compound action potentials in the field recording were observed and compared to the control. Results and Discussion: Compound action potentials show little or no synaptic input, increased delay and decreased amplitude with higher glucose concentration exposure as shown in Figure 2. The higher glucose concentration is thought to trigger the sorbitol pathway and induce oxidative stress in the neurons of the DRG, leading to apoptosis and axonal degeneration. Figure 2: Average traces representing compound action potentials recorded with and without high glucose treatment. The change in amplitude and latency are also quantified as bar plots. (n=10, * = p<0.05 and ***=p<0.001) Conclusion: Microengineered myelinated neural fiber tracts have morphology, physiology and, toxicity response analogous to a peripheral nerve. The platform could be used as an in-vitro model for neuropathy studies and drug development. Funding provided by Tulane University

Cerebrolysin improves sciatic nerve dysfunction in a mouse model of diabetic peripheral neuropathy.

To examine the effects of Cerebrolysin on the treatment of diabetic peripheral neuropathy, we first established a mouse model of type 2 diabetes mellitus by administering a high-glucose, high-fat diet and a single intraperitoneal injection of streptozotocin. Mice defined as diabetic in this model were then treated with 1.80, 5.39 or 8.98 mL/kg of Cerebrolysin via intraperitoneal injections for 10 consecutive days. Our results demonstrated that the number, diameter and area of myelinated nerve fibers increased in the sciatic nerves of these mice after administration of Cerebrolysin. The results of several behavioral tests showed that Cerebrolysin dose-dependently increased the slope angle in the inclined plane test (indicating an improved ability to maintain body position), prolonged tail-flick latency and foot-licking time (indicating enhanced sensitivity to thermal and chemical pain, respectively, and reduced pain thresholds), and increased an index of sciatic nerve function in diabetic mice compared with those behavioral results in untreated diabetic mice. Taken together, the anatomical and functional results suggest that Cerebrolysin ameliorated peripheral neuropathy in a mouse model of type 2 diabetes mellitus.

Ethanol Extracts from Toona sinensis Seeds Alleviates Diabetic Peripheral Neuropathy through Inhibiting Oxidative Stress and Regulating Growth Factor

Toona sinensis (A. Juss.) Roem has long been used in traditional Chinese medicine for the treatment of diabetic mellitus. The aim of this study was to investigate the effect of ethanol extracts of T. sinensis seeds on diabetic peripheral neuropathy and the preliminary mechanism. Male wistar rats were used for inducing diabetic peripheral neuropathy model, the variation of body weight; blood glucose, general status, thermal perception thresholds and sciatic nerve conduction velocity were used to confirm the diagnosis of diabetic peripheral neuropathy. After the treatment, the sciatic nerve conduction velocity was accelerated, the tailflick latency was reduced, the hyperglycemia and oxidative stress injury in sciatic nerve were ameliorated, contents of serum nerve growth factor β were up-regulated and tumor necrosis factor α, interleukin-6 were down-regulated significantly (P<0.05) compared with the model rats. This research demonstrates that ethanol extracts of T. sinensis seeds offers potential for intervening diabetic peripheral neuropathy and the protective mechanisms may be alleviating hyperglycemia, inhibiting oxidative stress and regulating growth factor and inflammatory cytokines.

An Introduction to the History and Controversies of Animal Models of Diabetic Neuropathy.

An Introduction to the History and Controversies of Animal Models of Diabetic Neuropathy.

Effect of levetiracetam versus gabapentin on peripheral neuropathy and sciatic degeneration in streptozotocin-diabetic mice: Influence on spinal microglia and astrocytes

Peripheral diabetic neuropathy develops in diabetic patients. The current study tested the antiallodynic and antihyperalgesic effects of the anticonvulsant drug, levetiracetam compared with the standard drug, gabapentin, in a model of streptozotocin-induced peripheral diabetic neuropathy. Male albino mice were injected intraperitoneally with streptozotocin (40mg/kg) for five consecutive days to induce type 1 diabetes mellitus. After development of peripheral diabetic neuropathy, mice were then treated orally with 10 doses of levetiracetam or gabapentin (or vehicle). The effect of multiple doses of levetiracetam on the histopathology of sciatic nerve and spinal cord was tested. Furthermore, the effect of levetiracetam on the spinal expression of microglia and astrocytes was examined in comparison with gabapentin. Results indicated that the highest dose of levetiracetam and all doses of gabapentin increased the withdrawal threshold in von Frey test. Furthermore, all doses of levetiracetam and gabapentin prolonged the reaction time exhibited by diabetic mice tested in hot plate test. Both drugs provided protection for the sciatic nerve and the spinal cord. In addition, levetiracetam (20 and 40mg/kg) decreased spinal immunostaining for CD11b (microglia marker) and glial fibrillary acidic protein (GFAP, astrocytes marker) however; the high dose of gabapentin (40mg/kg) reduced the spinal immunostaining for GFAP only. In conclusion, levetiracetam produced antiallodynic and antihyperalgesic effect in diabetic mice with favorable effects on sciatic nerve and spinal cord that were accompanied by downregulation of the spinal expression of microglia and astrocytes. Thus, levetiracetam may have promise in alleviating neuropathic pain in diabetic patients.

The Efficacy of Eslicarbazepine Acetate in Models of Trigeminal, Neuropathic, and Visceral Pain: The Involvement of 5-HT1B/1D Serotonergic and CB1/CB2 Cannabinoid Receptors.

Many clinical pain states that are difficult to treat share a common feature of sensitization of nociceptive pathways. Drugs that could normalize hyperexcitable neural activity (e.g., antiepileptic drugs) may be useful in relieving these pain states. Eslicarbazepine acetate (ESL) is a novel antiepileptic drug derived from carbamazepine/oxcarbazepine with a more favorable metabolic profile and potentially better tolerability. We examined the efficacy of ESL in models of inflammatory and neuropathic pain and the potential mechanism involved in its action. The antinociceptive effects of ESL were assessed in mice models of trigeminal (orofacial formalin test), neuropathic (streptozotocin-induced diabetic neuropathy model), and visceral pain (writhing test). The influence of 5-HT1B/1D serotonin receptor (GR 127935) and CB1 (AM251) and CB2 cannabinoid receptor (AM630) antagonists on the antinociceptive effect of ESL was tested in the model of trigeminal pain. ESL exhibited significant and dose-dependent antinociceptive effects in the second phase of the orofacial formalin test (P ≤ 0.011), in the tail-flick test in diabetic mice (P ≤ 0.013), and in the writhing test (P ≤ 0.003). GR 127935 (P ≤ 0.038) and AM251 and AM630 (P ≤ 0.013 for both antagonists) significantly inhibited the antinociceptive effect of ESL in a dose-related manner. ESL exhibited efficacy in models of trigeminal, neuropathic, and visceral pain. In the trigeminal pain model, the antinociceptive effect of ESL is, at least in part, mediated by 5-HT1B/1D serotonin and CB1/CB2 cannabinoid receptors. This study indicates that ESL could be useful in the clinical treatment of inflammatory and neuropathic pain.

Anti-oxidative and anti-inflammatory effects of cinnamaldehyde on protecting high glucose-induced damage in cultured dorsal root ganglion neurons of rats.

To examine the mechanism underlying the beneficial role of cinnamaldehyde on oxidative damage and apoptosis in high glucose (HG)-induced dorsal root ganglion (DRG) neurons in vitro. HG-treated DRG neurons were developed as an in vitro model of diabetic neuropathy. The neurons were randomly divided into five groups: the control group, the HG group and the HG groups treated with 25, 50 and 100 nmol/L cinnamaldehyde, respectively. Cell viability was examined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and apoptosis rate was evaluated by the in situ TdT-mediated dUTP nick end labeling (TUNEL) assay. The intracellular level of reactive oxygen species (ROS) was measured with flow cytometry. Expression of nuclear factor-kappa B (NF-κB), inhibitor of κB (IκB), phosphorylated IκB (p-IκB), tumor necrosis factor (TNF)-α, interleukin-6 (IL-6) and caspase-3 were determined by western blotting and real-time quantitative reverse transcription polymerase chain reaction (RT-PCR). Expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and hemeoxygenase-1 (HO-1) were also measured by western blotting. Cinnamaldehyde reduced HG-induced loss of viability, apoptosis and intracellular generation of ROS in the DRG neurons via inhibiting NF-κB activity. The western blot assay results showed that the HG-induced elevated expressions of NF-κB, IκB and p-IκB were remarkably reduced by cinnamaldehyde treatment in a dose-dependent manner (P <0.01). The HG-induced over-expression of NF-κB p65 mRNA was remarkably attenuated after cinnamaldehyde treatment in a dose-dependent manner (P <0.01). However, the expressions of Nrf2 and HO-1 were not upregulated. Treatment with cinnamaldehyde not only attenuated caspase-3 activation and the caspase cleavage cascade in DRG neurons, but also lowered the elevated IL-6, TNF-α, cyclo-oxygenase and inducible nitric oxide synthase levels, indicating a reduction in inflammatory damage. Cinnamaldehyde protected DRG neurons from the deleterious effects of HG through inactivation of NF-κB pathway but not through activation of Nrf2/HO-1. And thus cinnamaldehyde may have potential application as a treatment for DPN.

Gender-specific differences in diabetic neuropathy in BTBR ob/ob mice

AimsTo identify a female mouse model of diabetic peripheral neuropathy (DPN), we characterized DPN in female BTBR ob/ob mice and compared their phenotype to non-diabetic and gender-matched controls. We also identified dysregulated genes and pathways in sciatic nerve (SCN) and dorsal root ganglia (DRG) of female BTBR ob/ob mice to determine potential DPN mechanisms. MethodsTerminal neuropathy phenotyping consisted of examining latency to heat stimuli, sciatic motor and sural sensory nerve conduction velocities (NCV), and intraepidermal nerve fiber (IENF) density. For gene expression profiling, DRG and SCN were dissected, RNA was isolated and processed using microarray technology and differentially expressed genes were identified. ResultsSimilar motor and sensory NCV deficits were observed in male and female BTBR ob/ob mice at study termination; however, IENF density was greater in female ob/ob mice than their male counterparts. Male and female ob/ob mice exhibited similar weight gain, hyperglycemia, and hyperinsulinemia compared to non-diabetic controls, although triglycerides were elevated more so in males than in females. Transcriptional profiling of nerve tissue from female mice identified dysregulation of pathways related to inflammation. ConclusionsSimilar to males, female BTBR ob/ob mice display robust DPN, and pathways related to inflammation are dysregulated in peripheral nerve.

2015 4thTERMIS World CongressBoston, MassachusettsSeptember 8–11, 2015

2015 4^thTERMIS World CongressBoston, MassachusettsSeptember 8–11, 2015

The furoxan nitric oxide donor, PRG150, evokes dose-dependent analgesia in a rat model of painful diabetic neuropathy.

Painful diabetic neuropathy (PDN) is a type of peripheral neuropathic pain that is often intractable. Elevated nitric oxide (NO) from neuronal and non-neuronal sources in the somatosensory system is implicated in the pathobiology of peripheral neuropathic pain. However, in diabetes, nitrergic nerve degeneration to deplete NO bioactivity appears causal in the pathogenesis of irreversible autonomic neuropathy, another long term complication of diabetes. Hence, this study hypothesized that progressive NO depletion may underpin the pathobiology of PDN and that NO donors may alleviate PDN. Diabetes was induced in rats with intravenous streptozotocin (STZ) at 70mg/kg and confirmed if blood glucose levels (BGLs) on day 10 post-STZ were ≥15mmol/L. Analgesic efficacy of subcutaneous (s.c.) bolus doses of the furoxan NO donor, PRG150 was assessed in the STZ-diabetic rat model of PDN at 10-, 14- and 24-weeks post-STZ relative to the sydnominine NO donor, SIN-1 and its prodrug, molsidomine. PRG150 produced dose-dependent analgesia in STZ-diabetic rats whereas SIN-1 and molsidomine evoked neuro-excitatory side-effects, but not analgesia. The 1000-fold larger doses of PRG150 needed to produce analgesia at 14- and 24-weeks (800pmol/kg) c.f. 10-weeks (8fmol/kg) post-STZ in rats, suggest that progressive NO depletion is also causal in PDN. Importantly, doses of PRG150 up to 10000 fold higher than the analgesic dose did not produce hypotension in rats. The 50-fold greater release of NO by SIN-1 c.f. PRG150 invitro, may underpin the neuro-excitatory rather than analgesic effects of SIN-1/molsidomine. PRG150 is worthy of further investigation as a potential novel analgesic for PDN.

Histopathological and behavioral evaluations of the effects of crocin, safranal and insulin on diabetic peripheral neuropathy in rats.

Crocin and safranal, the major constituents of saffron, exert neuroprotective effects. In the present study, we investigated the effects of crocin and safranal (alone or in combination with insulin) on peripheral neuropathy in diabetic rats. Diabetes was induced by intraperitoneal (i.p.) injection of 60 mg/kg of streptozotocin (STZ) and confirmed by blood glucose level higher than 250 mg/dl. After confirmation of diabetes, crocin (30 mg/kg, i.p.), safranal (1 mg/kg, i.p.) (alone or in combination with insulin) and insulin (5 IU/kg, s.c.) were administered for eight weeks. Neuropathic pain was evaluated using acetone drop test. Histopathological changes of sciatic nerve were evaluated using light microscope. Blood glucose levels and sciatic nerve malondialdehyde (MDA) contents were also measured. STZ caused cold allodynia, edema and degenerative changes of sciatic nerve, hyperglycemia and an elevation of sciatic nerve MDA levels. Crocin, safranal and insulin improved STZ-induced behavioral, histopathological and biochemical changes. Combined treatments produced more documented improving effects. The results of the present study showed neuroprotective effects of crocin, safranal and insulin in a rat model of diabetic neuropathy. In addition, crocin and safranal enhanced the neuroprotective effect of insulin. The neuroprotective effects of theses chemical compounds could be associated with their anti-hyperglycemic and antioxidant properties.

Streptozotocin Stimulates the Ion Channel TRPA1 Directly: INVOLVEMENT OF PEROXYNITRITE

Streptozotocin (STZ)-induced diabetes is the most commonly used animal model of diabetes. Here, we have demonstrated that intraplantar injections of low dose STZ evoked acute polymodal hypersensitivities in mice. These hypersensitivities were inhibited by a TRPA1 antagonist and were absent in TRPA1-null mice. In wild type mice, systemic STZ treatment (180 mg/kg) evoked a loss of cold and mechanical sensitivity within an hour of injection, which lasted for at least 10 days. In contrast, Trpa1(-/-) mice developed mechanical, cold, and heat hypersensitivity 24 h after STZ. The TRPA1-dependent sensory loss produced by STZ occurs before the onset of diabetes and may thus not be readily distinguished from the similar sensory abnormalities produced by the ensuing diabetic neuropathy. In vitro, STZ activated TRPA1 in isolated sensory neurons, TRPA1 cell lines, and membrane patches. Mass spectrometry studies revealed that STZ oxidizes TRPA1 cysteines to disulfides and sulfenic acids. Furthermore, incubation of tyrosine with STZ resulted in formation of dityrosine, suggesting formation of peroxynitrite. Functional analysis of TRPA1 mutants showed that cysteine residues that were oxidized by STZ were important for TRPA1 responsiveness to STZ. Our results have identified oxidation of TRPA1 cysteine residues, most likely by peroxynitrite, as a novel mechanism of action of STZ. Direct stimulation of TRPA1 complicates the interpretation of results from STZ models of diabetic sensory neuropathy and strongly argues that more refined models of diabetic neuropathy should replace the use of STZ.

Antihyperalgesic effects of ProTx-II, a Nav1.7 antagonist, and A803467, a Nav1.8 antagonist, in diabetic mice.

The present study investigated the effects of intrathecal administration of ProTx-II (tarantula venom peptide) and A803467 (5-[4-chloro-phenyl]-furan-2-carboxylic acid [3,5-dimethoxy-phenyl]-amide), selective Nav1.7 and Nav1.8 antagonists, respectively, on thermal hyperalgesia in a painful diabetic neuropathy model of mice. Intrathecal administration of ProTx-II at doses from 0.04 to 4 ng to diabetic mice dose-dependently and significantly increased the tail-flick latency. Intrathecal administration of A803467 at doses from 10 to 100 ng to diabetic mice also dose-dependently and significantly increased the tail-flick latency. However, intrathecal administration of either ProTx-II (4 ng) or A803467 (100 ng) had no effect on the tail-flick latency in nondiabetic mice. The expression of either the Nav1.7 or Nav1.8 sodium channel protein in the dorsal root ganglion in diabetic mice was not different from that in nondiabetic mice. The present results suggest that ProTx-II and A803467, highly selective blockers of Nav1.7 and Nav1.8 sodium channels, respectively, in the spinal cord, can have antihyperalgesic effects in diabetic mice.

Levetiracetam synergises with common analgesics in producing antinociception in a mouse model of painful diabetic neuropathy

Painful diabetic neuropathy is difficult to treat. Single analgesics often have insufficient efficacy and poor tolerability. Combination therapy may therefore be of particular benefit, because it might provide optimal analgesia with fewer adverse effects. This study aimed to examine the type of interaction between levetiracetam, a novel anticonvulsant with analgesic properties, and commonly used analgesics (ibuprofen, aspirin and paracetamol) in a mouse model of painful diabetic neuropathy. Diabetes was induced in C57BL/6 mice with a single high dose of streptozotocin, applied intraperitoneally (150mg/kg). Thermal (tail-flick test) and mechanical (electronic von Frey test) nociceptive thresholds were measured before and three weeks after diabetes induction. The antinociceptive effects of orally administered levetiracetam, analgesics, and their combinations were examined in diabetic mice that developed thermal/mechanical hypersensitivity. In combination experiments, the drugs were co-administered in fixed-dose fractions of single drug ED50 and the type of interaction was determined by isobolographic analysis. Levetiracetam (10–100mg/kg), ibuprofen (2–50mg/kg), aspirin (5–75mg/kg), paracetamol (5–100mg/kg), and levetiracetam-analgesic combinations produced significant, dose-dependent antinociceptive effects in diabetic mice in both tests. In the tail-flick test, isobolographic analysis revealed 15-, and 19-fold reduction of doses of both drugs in the combination of levetiracetam with aspirin/ibuprofen, and paracetamol, respectively. In the von Frey test, approximately 7- and 9-fold reduction of doses of both drugs was detected in levetiracetam-ibuprofen and levetiracetam-aspirin/levetiracetam-paracetamol combinations, respectively. These results show synergism between levetiracetam and ibuprofen/aspirin/paracetamol in a model of painful diabetic neuropathy and might provide a useful approach to the treatment of patients suffering from painful diabetic neuropathy.

Small organic molecule disruptors of Cav3.2 - USP5 interactions reverse inflammatory and neuropathic pain.

BackgroundCav3.2 channels facilitate nociceptive transmission and are upregulated in DRG neurons in response to nerve injury or peripheral inflammation. We reported that this enhancement of Cav3.2 currents in afferent neurons is mediated by deubiquitination of the channels by the deubiquitinase USP5, and that disrupting USP5/Cav3.2 channel interactions protected from inflammatory and neuropathic pain.ResultsHere we describe the development of a small molecule screening assay for USP5-Cav3.2 disruptors, and report on two hits of a ~5000 compound screen - suramin and the flavonoid gossypetin. In mouse models of inflammatory pain and neuropathic pain, both suramin and gossypetin produced dose-dependent and long-lasting mechanical anti-hyperalgesia that was abolished or greatly attenuated in Cav3.2 null mice. Suramin and Cav3.2/USP5 Tat-disruptor peptides were also tested in models of diabetic neuropathy and visceral pain, and provided remarkable protection.ConclusionsOverall, our findings provide proof of concept for a new class of analgesics that target T-type channel deubiquitination.Electronic supplementary materialThe online version of this article (doi:10.1186/s12990-015-0011-8) contains supplementary material, which is available to authorized users.

Aqueous extract of Zizyphus jujuba fruit attenuates glucose induced neurotoxicity in an in vitro model of diabetic neuropathy.

The neuroprotective effect of fruit aqueous extract of Ziziphus jujuba Lam on glucose-induced neurotoxicity in PC12 cells as an appropriate in vitro model of diabetic neuropathy was investigated. Cell viability was determined by the MTT assay. Cellular reactive oxygen species (ROS) generation was measured by DCFH-DA analysis. Cleaved caspase-3, a biochemical parameter of cellular apoptosis, was measured by western blot analysis. Our data showed that a 4-fold elevation in glucose levels within the medium significantly reduced cell viability, increased intracellular ROS and caspase-3 activation in PC12 cells after 24 hr. Incubation of the high glucose medium cells with 300-μg/ml Z. jujuba fruit (ZJF) extract decreased the high glucose-induced cell toxicity and prevented caspase-3 activation and excited ROS generation. Thus, we concluded that the aqueous extract of Z. jujuba protects against hyperglycaemia-induced cellular toxicity. This could be associated with the prevention of ROS generation and neural apoptosis. Moreover, the results suggest that the ZJF has a therapeutic potential to attenuate diabetes complications such as neuropathy.