Dorsal Root Ganglia Of Diabetic Rats Research Articles

Salidroside alleviates diabetic neuropathic pain through regulation of the AMPK-NLRP3 inflammasome axis

High glucose (HG)-induced nucleotide-binding and oligomerization (NACHT) domain, leucine-rich repeat (LRR), and pyrin domain (PYD)-containing protein 3 (NLRP3) inflammasome activation leads to diabetic neuropathic pain. We recently showed that salidroside could suppress NLRP3 inflammasome activation in hepatocytes exposed to HG. The aim of this study was to evaluate the analgesic effect of salidroside on diabetic rats and to explore its underlying mechanisms. Rat models with diabetic neuropathic pain were induced by high-fat diet feeding combined with low dose streptozotocin injections. Doses of salidroside at 50 and 100 mg.kg−1.day−1 were administered by gavage to diabetic rats for 6 weeks. Mechanical allodynia test, thermal hyperalgesia test and biochemical analysis were performed to evaluate therapeutic effects. Primary dorsal root ganglion (DRG) cells exposed to HG at 45 mM were used to further study the effects of salidroside on the AMP-activated protein kinase (AMPK)-NLRP3 inflammasome axis and insulin sensitivity in vitro. Salidroside administration improved hyperglycemia, ameliorated insulin resistance, and alleviated neuropathic pain in diabetic rats. Moreover, salidroside induced AMPK activation and suppressed NLRP3 inflammasome activation in the DRGs of diabetic rats. In addition, salidroside treatment relieved oxidative stress, improved insulin sensitivity and regulated the AMPK-NLRP3 inflammasome axis in HG-treated DRGs in vitro. Furthermore, AMPK inhibition in vivo or AMPK silencing in vitro abolished the beneficial effects of salidroside on diabetic neuropathic pain. Together, these results indicate that salidroside alleviates diabetic neuropathic pain through its regulation of the AMPK-NLRP3 inflammasome axis in DRGs.

Quercetin relieved diabetic neuropathic pain by inhibiting upregulated P2X4 receptor in dorsal root ganglia.

The upregulation of nociceptive ion channels expressed in dorsal root ganglia (DRG) contributes to the development and retaining of diabetic pain symptoms. The flavonoid quercetin (3,3',4',5,7-pentahydroxyflavone) is a component extracted from various fruits and vegetables and exerts anti-inflammatory, analgesic, anticarcinogenic, antiulcer, and antihypertensive effects. However, the exact mechanism underlying quercetin's analgesic action remains poorly understood. The aim of this study was to investigate the effects of quercetin on diabetic neuropathic pain related to the P2X4 receptor in the DRG of type 2 diabetic rat model. Our data showed that both mechanical withdrawal threshold and thermal withdrawal latency in diabetic rats treated with quercetin were higher compared with those in untreated diabetic rats. The expression levels of P2X4 messenger RNA and protein in the DRG of diabetic rats were increased compared with the control rats, while quercetin treatment significantly inhibited such enhanced P2X4 expression in diabetic rats. The satellite glial cells (SGCs) enwrap the neuronal soma in the DRG. Quercetin treatment also lowered the elevated coexpression of P2X4 and glial fibrillary acidic protein (a marker of SGCs) and decreased the upregulation of phosphorylated p38 mitogen-activated protein kinase (p38MAPK) in the DRG of diabetic rats. Quercetin significantly reduced the P2X4 agonist adenosine triphosphate-activated currents in HEK293 cells transfected with P2X4 receptors. Thus, our data demonstrate that quercetin may decrease the upregulation of the P2X4 receptor in DRG SGCs, and consequently inhibit P2X4 receptor-mediated p38MAPK activation to relieve the mechanical and thermal hyperalgesia in diabetic rats.

Tanshinone IIA Improves Painful Diabetic Neuropathy by Suppressing the Expression and Activity of Voltage-Gated Sodium Channel in Rat Dorsal Root Ganglia.

Painful diabetic neuropathy (PDN) is one of the intractable complications of diabetes mellitus, which manifest as exaggerated pain perception. Previous studies showed that Tanshinone IIA (TIIA), one of the major bioactive extracts of Salvia miltiorrhiza Bunge, have obvious analgesic effect on different types of pain process, and the underlying analgesic mechanisms are not fully understood. The present study combined the behavioral, electrophysiological and biochemical methods to elucidate the analgesic mechanism of TIIA, using streptozotocin (STZ)-induced PDN rat models. Intraperitoneal injection (i.p.) of TIIA for 3 weeks in PDN rats significantly improved mechanical allodynia and thermal hyperalgesia. Patch clamp recordings showed that the excitability of dorsal root ganglion (DRG) nociceptive neuron was increased in diabetic state, and TIIA treatment effectively recovered the subnormality, which was achieved by preventing augments of both Tetrodotoxin-sensitive (TTX-resistant) and Tetrodotoxin-sensitive (TTX-S) sodium currents. Further, the protein expressions of voltage-gated sodium channels (VGSCs) α-subunits Nav1.3, Nav1.7 and Nav1.9 increased in DRG of diabetic rats and were normalized by TIIA application. In conclusion, this study provides evidence that the TIIA attenuated PDN by effecting VGSCs activities and expressions, indicating that the TIIA could be a promising agent for PDN treatment.

Insulin prevents aberrant mitochondrial phenotype in sensory neurons of type 1 diabetic rats

Diabetic neuropathy affects approximately 50% of diabetic patients. Down-regulation of mitochondrial gene expression and function has been reported in both human tissues and in dorsal root ganglia (DRG) from animal models of type 1 and type 2 diabetes. We hypothesized that loss of direct insulin signaling in diabetes contributes to loss of mitochondrial function in DRG neurons and to development of neuropathy. Sensory neurons obtained from age-matched adult control or streptozotocin (STZ)-induced type 1 diabetic rats were cultured with or without insulin before determining mitochondrial respiration and expression of mitochondrial respiratory chain and insulin signaling-linked proteins. For in vivo studies age-matched control rats and diabetic rats with or without trace insulin supplementation were maintained for 5months before DRG were analyzed for respiratory chain gene expression and cytochrome c oxidase activity. Insulin (10nM) significantly (P<0.05) increased phosphorylation of Akt and P70S6K by 4-fold and neurite outgrowth by 2-fold in DRG cultures derived from adult control rats. Insulin also augmented the levels of selective mitochondrial respiratory chain proteins and mitochondrial bioenergetics parameters in DRG cultures from control and diabetic rats, with spare respiratory capacity increased by up to 3-fold (P<0.05). Insulin-treated diabetic animals exhibited improved thermal sensitivity in the hind paw and had increased dermal nerve density compared to untreated diabetic rats, despite no effect on blood glucose levels. In DRG of diabetic rats there was suppressed expression of mitochondrial respiratory chain proteins and cytochrome c oxidase activity that was corrected by insulin therapy. Insulin elevates mitochondrial respiratory chain protein expression and function in sensory neurons and this is associated with enhanced neurite outgrowth and protection against indices of neuropathy.

Alpha-lipoic Acid suppresses P2X receptor activities and visceral hypersensitivity to colorectal distention in diabetic rats

The present study was designed to investigate the roles of P2X3 receptors in dorsal root ganglion (DRG) neurons in colonic hypersensitivity and the effects of alpha-lipoic acid (ALA) on P2X3 receptor activity and colonic hypersensitivity of diabetic rats. Streptozotocin (STZ) was used to induce diabetic model. Abdominal withdrawal reflex (AWR) responding to colorectal distention (CRD) was recorded as colonic sensitivity. ATP-induced current density of colon-specific DRG (T13-L2 DRGs) neurons was measured with whole-cell patch clamp. The expression of P2X3Rs of T13-L2 DRGs was measured by western blot analysis. The results showed that AWR scores significantly increased after STZ injection. P2X3R expression and ATP current density of T13-L2 DRG neurons were enhanced in diabetic rats. Intraperitoneal injection with ALA once a day for 1 week remarkably reduced P2X3R expression and ATP current density in diabetic rats. Importantly, ALA treatment attenuated colonic hypersensitivity in diabetic rats. Our data suggest that STZ injection increases expression and function of P2X3 receptors of colon-specific DRG neurons, thus contributing to colonic hypersensitivity in diabetic rats. Administration of ALA attenuates diabetic colonic hypersensitivity, which is most likely mediated by suppressing expression and function of P2X3 receptors in DRGs of diabetic rats.

Exogenous brain-derived neurotrophic factor relieves pain symptoms of diabetic rats by reducing excitability of dorsal root ganglion neurons

Diabetic peripheral neuropathy (DPN) is a common complication of diabetes lacking of effective treatments. Enhanced excitability of dorsal root ganglion (DRG) neuron plays a crucial role in the progression of diabetic neuropathic hyperalgesia. Brain-derived neurotrophic factor (BDNF) is known as a neuromodulator of nociception, but whether and how BDNF modulates the excitability of DRG neurons in the development of DPN remain to be clarified. This study investigated the role of exogenous BDNF and its high-affinity tropomyosin receptor kinase B (TrkB) in rats with streptozotocin-induced diabetic neuropathic pain. The results showed that continued intrathecal administration of BDNF to diabetic rats dramatically alleviated mechanical and thermal hyperalgesia, as well as inhibited hyperexcitability of DRG neurons. These effects were blocked by pretreatment with TrkB Fc (a synthetic fusion protein consisting of the extracellular ligand-binding domain of the TrkB receptor). The expression of BDNF and TrkB was upregulated in the DRG of diabetic rats. Intrathecal administration of BDNF did not affect this upregulation. These data provide novel information that exogenous BDNF relieved pain symptoms of diabetic rats by reducing hyperexcitability of DRG neurons and might be the potential treatment of painful diabetic neuropathy.

Dietary fish oil inhibits mechanical allodynia and thermal hyperalgesia in diabetic rats by blocking nuclear factor-κB-mediated inflammatory pathways

One of the most common complications of early-onset diabetes mellitus is peripheral diabetic neuropathy, which is manifested either by loss of nociception or by allodynia and hyperalgesia. Diabetes mellitus is a common metabolic disease in human beings with characteristic symptoms of hyperglycemia, chronic inflammation and insulin resistance. Dietary fatty acids, especially polyunsaturated fatty acids, have been shown anti-inflammatory role in various experimental conditions. The present study investigated the effects of fish oil supplementation on the inflammation in the dorsal root ganglion (DRG) of streptozotocin (STZ)-induced diabetes rats. The effects of diabetes and fish oil treatment on the allodynia and hyperalgesia were also evaluated. Dietary fish oil effectively attenuated both allodynia and hyperalgesia induce by STZ injection. Along with the behavioral findings, DRG from fish oil-treated diabetic rats displayed a decrease in inflammatory cytokines and the expression of nuclear factor-κB (NF-κB) compared with untreated diabetic rats. Fish oil supplementation also increased the phosphorylation of AKT in DRG of diabetic rats. These results suggested that dietary fish oil-inhibited allodynia and hyperalgesia in diabetic rats may stem from its anti-inflammatory potential by regulating NF-κB and AKT. Fish oil might be useful as an adjuvant therapy for the prevention and treatment of diabetic complications.

Virus-Mediated Knockdown of Nav1.3 in Dorsal Root Ganglia of STZ-Induced Diabetic Rats Alleviates Tactile Allodynia.

Diabetic neuropathic pain affects a substantial number of people and represents a major public health problem. Available clinical treatments for diabetic neuropathic pain remain only partially effective and many of these treatments carry the burden of side effects or the risk of dependence. The misexpression of sodium channels within nociceptive neurons contributes to abnormal electrical activity associated with neuropathic pain. Voltage-gated sodium channel Nav1.3 produces tetrodotoxin-sensitive sodium currents with rapid repriming kinetics and has been shown to contribute to neuronal hyperexcitability and ectopic firing in injured neurons. Suppression of Nav1.3 activity can attenuate neuropathic pain induced by peripheral nerve injury. Previous studies have shown that expression of Nav1.3 is upregulated in dorsal root ganglion (DRG) neurons of diabetic rats that exhibit neuropathic pain. Here, we hypothesized that viral-mediated knockdown of Nav1.3 in painful diabetic neuropathy would reduce neuropathic pain. We used a validated recombinant adeno-associated virus (AAV)-shRNA-Nav1.3 vector to knockdown expression of Nav1.3, via a clinically applicable intrathecal injection method. Three weeks following vector administration, we observed a significant rate of transduction in DRGs of diabetic rats that concomitantly reduced neuronal excitability of dorsal horn neurons and reduced behavioral evidence of tactile allodynia. Taken together, these findings offer a novel gene therapy approach for addressing chronic diabetic neuropathic pain.

Expression of nerve growth factor in lumbar dorsal root ganglia and bladder of streptozotocin-induced diabetic rats

Objective To investigate the expression of nerve growth factor (NGF) in lumbar dorsal root ganglia (DRG) and bladder of streptozotocin-induced diabetic rats before and after treatment.Methods Thirty male Sprague-Dawley (SD) rats were divided into five groups randomly,including 6 in control group,24 in diabetic cystopathy (DC) group.The rat DC models were made throught injection of STZ.The rats in DC group were divided into four subgroups randomly (n =6 each),including DC group,NGF group,RI group,and NRG + RI group.The diabetic rats were treated with NGF and insulin for 4 weeks.NGF was detected in the bladder and DRG by using immunohistochemical staining.Results The blood sugar level in DC group was significantly higher than that in the control group [(30.90 ± 2.33) vs.(4.30 ±0.18) mmol/L (t =30.33,P ＜ 0.01)].The NGF levels in both bladder and DRG of diabetic rats treated with NGF and insulin were significantly increased (0.1492 ± 0.0046 vs.0.1066 ± 0.0055,P＜0.05; 0.1667 ±0.0065 vs.0.1174 ±0.0050,P＜0.05).Conclusion The aberrant expression of NGF in both bladder and DRG of diabetic rats may play an important role,and using exogenous NGF may help to alleviate the pathological changes and improve the function of rats with DC. Key words: Diabetes mellitus; Detrusor; Dorsal root ganglion; Nerve growth factor

Early potential impairment of renal sensory nerves in streptozotocin-induced diabetic rats: role of neurokinin receptors

Electrophysiological studies in the mammalian kidney have identified two major classes of sensory receptors of the afferent renal nerves; chemoreceptors (CR) and mechanoreceptors (MR). The localization of calcitonin gene-related peptide (CGRP) and substance P (SP) in these renal pelvic sensory neurons provides an anatomical basis for a possible functional interaction between the two neuropeptides and SP receptor. The present study was performed to examine the possible changes in the responsiveness of renal sensory SP and CGRP receptors in rats with streptozotocin (STZ)-induced diabetes mellitus. Due to the crucial role of renal pelvic SP and CGRP receptors in the activation of renal sensory neurons by various stimuli, we examined whether the responsiveness of MR or CR activation and the dorsal root ganglia content of neuropeptides and neurokinin 1 receptors (NK(1)R) were altered in diabetic rats compared with non-diabetic rats. Afferent renal nerve activity (ARNA) was recorded from the peripheral portion of the cut end of one renal nerve branch placed on a bipolar silver wire electrode. T(13) dorsal root ganglia (DRG) immunoreactivity was performed to NK(1)R, SP and CGRP. The results of the current study confirmed that the stimulation of renal MR and CR elicited a renorenal reflex response, and that the renal pelvic administration of SP and CGRP increased ipsilateral ARNA and contralateral urinary sodium excretion with no changes in arterial pressure. We also found a decrease in NK(1)R expression followed by an increase in SP and CGRP levels in the DRG of diabetic rats. The ARNA response, produced by renal pelvic MR and CR stimulation, was found to be significantly attenuated in the STZ-induced diabetic model. Conclusions. These data may indicate a compensatory synthesis and/or abnormal axonal delivery of neurokinins from the cell body to synaptic portions of the neuron as the underlying reason for attenuated ARNA in renal sensory neurons of diabetic rats.

Reduced expression of endothelin B receptors and mechanical hyperalgesia in experimental chronic diabetes

Diabetic neuropathy is one of the major complications of diabetes mellitus. Small nerve fibers degenerate early in the disease, leading to symptoms ranging from hyperalgesia to loss of pain and temperature sensation. However, the cellular and molecular mechanisms responsible for abnormal pain perception in diabetes have not been identified. Both type-A and type-B endothelin receptors (ETAR and ETBR, respectively) are present in sensory nerves and appear to regulate neuropathic and inflammatory pain. In this study, we compared the expression of endothelin receptors and nociceptive responses in normal and experimentally diabetic rats. Diabetic animals exhibited both an increase in the withdrawal responses to high threshold stimuli (mechanical hyperalgesia) and to light touch stimuli (tactile allodynia). Immunohistochemical and Western blot analysis revealed that diabetic rats have significantly reduced expression of ETBR in sciatic nerves, while no changes were observed in dorsal root ganglia (DRG). In contrast, the expression of ETAR in either sciatic nerves or DRG of diabetic rats was not altered. Importantly, ETBR-deficient transgenic rats showed alterations in pain perception similar to those observed in diabetic rats. These results suggest that changes in the expression of ETBR in peripheral nerve may contribute to the development of mechanical hyperalgesia and tactile allodynia in chronic diabetes.

Nerve growth factor (NGF) and diabetic neuropathy in the rat: morphological investigations of the sural nerve, dorsal root ganglion, and spinal cord.

A number of functions for nerve growth factor (NGF) have been described over the past years, including its role for neuronal function and regeneration during toxic or metabolic neuropathies. In order to further assess the effects of NGF on the somatosensory system in diabetic neuropathy, the sural nerve, dorsal root ganglia (DRG), and dorsal horn of the spinal cord were investigated by morphological and quantitative methods in rats after 12 weeks of uncontrolled streptozotocin-induced diabetes mellitus. The results from our study suggest a twofold effect of NGF: (1) In sural nerve treatment with NGF (0.1 or 0.5 mg/kg) for 12 weeks was able to reverse distinct diabetes-related alterations in myelinated nerve fiber morphology, such as myelin thickness. These changes occurred in the entire myelinated population of sensory nerves and were not restricted to nociceptive nerve fibers. (2) The NGF effect on neurotransmitters of the sensory, nociceptive system was reflected by increased CGRP and substance P content in the DRG and in the dorsal horn of the spinal cord. No change of trkA receptor immunostaining was seen in DRGs of diabetic rats; however, a reduction of trkA immunoreactivity of DRG neurons was noted after long-term NGF treatment of healthy controls. The data demonstrate that NGF regulates a number of neuronal parameters along peripheral and central parts of the somatosensory pathway in the adult. This neurotrophic support may be essential for inducing functionally significant regenerative mechanisms in diabetic neuropathy.