Synapses In Spinal Dorsal Horn Research Articles

The Surface Amine Group of Ultrasmall Magnetic Iron Oxide Nanoparticles Produce Analgesia in the Spinal Cord and Decrease Long-Term Potentiation.

Our previous studies have revealed the ultrasmall superparamagnetic iron oxide in the amine group USPIO-101 has an analgesic effect on inflammatory pain. Here, we further investigated its effect on the spinal cord and brain via electrophysiological and molecular methods. We used a mouse inflammatory pain model, induced by complete Freund’s adjuvant (CFA), and measured pain thresholds via von Frey methods. We also investigated the effects of USPIO-101 via an extracellular electrophysiological recording at the spinal dorsal horn synapses and hippocampal Schaffer collateral-CA1 synapses, respectively. The mRNA expression of pro-inflammatory cytokines was detected by quantitative real-time polymerase chain reaction (RT-qPCR). Our results showed intrathecal USPIO-101 produces similar analgesic behavior in mice with chronic inflammatory pain via intrathecal or intraplantar administration. The potentiated low-frequency stimulation-induced spinal cord long-term potentiation (LTP) at the spinal cord superficial dorsal horn synapses could decrease via USPIO-101 in mice with chronic inflammatory pain. However, the mRNA expression of cyclooxygenase-2 was enhanced with lipopolysaccharide (LPS) stimulation in microglial cells, and we also found USPIO-101 at 30 µg/mL could decrease the magnitude of hippocampal LTP. These findings revealed that intrathecal USPIO-101 presented an analgesia effect at the spinal cord level, but had neurotoxicity risk at higher doses.

Mechanisms for therapeutic effect of bulleyaconitine A on chronic pain.

Bulleyaconitine A, a diterpenoid alkaloid isolated from Aconitum bulleyanum plants, has been used for the treatment of chronic pain in China since 1985. Clinical studies show that the oral administration of bulleyaconitine A is effective for treating different kinds of chronic pain, including back pain, joint pain, and neuropathic pain with minimal side effect in human patients. The experimental studies have revealed that bulleyaconitine A at therapeutic doses potently inhibits the peripheral sensitization and central sensitization that underlie chronic pain and has no effect on acute pain. Bulleyaconitine A preferably blocks tetrodotoxin-sensitive voltage-gated sodium channels in dorsal root ganglion neurons by inhibition of protein kinase C, and the effect is around 600 times more potent in neuropathic animals than in naïve ones. Bulleyaconitine A at 5 nM inhibits the hypersensitivity of dorsal root ganglion neurons in neuropathic rats but has no effect on excitability of dorsal root ganglion neurons in sham group. Bulleyaconitine A inhibits long-term potentiation at C-fiber synapses in spinal dorsal horn, a synaptic model of pathological pain, preferably in neuropathic pain rats over naïve rats. The following mechanisms may underlie the selective effect of bulleyaconitine A on chronic pain. (1) In neuropathic conditions, protein kinase C and voltage-gated sodium channels in dorsal root ganglion neurons are upregulated, which enhances bulleyaconitine A’s effect. (2) Bulleyaconitine A use-dependently blocks voltage-gated sodium channels and therefore inhibits the ectopic discharges that are important for neuropathic pain. (3) Bulleyaconitine A is shown to inhibit neuropathic pain by the modulation of spinal microglia, which are involved in the chronic pain but not in acute (nociceptive) pain. Moreover, bulleyaconitine A facilitates the anesthetic effect of morphine and inhibits morphine tolerance in rats. Together, bulleyaconitine A is able to inhibit chronic pain by targeting at multiple molecules. Further clinical and experimental studies are needed for evaluating the efficacy of bulleyaconitine A in different forms of chronic pain in patients and for exploring the underlying mechanisms.

Stereological study on the number of synapses in the rat spinal dorsal horn with painful diabetic neuropathy induced by streptozotocin.

Our previous studies showed that direct injury to the sciatic nerve (chronic constriction injury or axotomy) is associated with a numerical increase in synaptic number in the rat spinal dorsal horn. The aim of this study was to determine whether painful diabetic neuropathy (PDN) was also associated with numerical changes in the synaptic or neuronal numbers in the spinal dorsal horn. Overall, 17 adult SD rats were allocated randomly into the control group (n=5) and the streptozotocin (STZ) group (n=12). STZ was injected intraperitoneally to induce diabetes. In the STZ group, seven rats (STZ-H) showed hyperglycemia (fasting blood glucose >11.1 mM) and the rest of the five rats (STZ-N) did not. Rats were fed and observed for 28 days after hyperglycemia. Two of the seven STZ-H rats died of infection during the observation period. Body weight and paw withdraw threshold (PWT) decreased in the rest of the five STZ-H rats. Twenty-eight days after hyperglycemia, the L5 segment of the spinal cord was removed; paraffin-embedded sections were prepared and stained with Nissl's method and synaptophysin immunohistochemistry, respectively. The optical dissector (a stereological technique) was used to estimate the numbers of neurons and synapses in the spinal dorsal horn. Compared with the control group, the synaptic number and ratio between the numbers of synapses and neurons in the L5 segment of the spinal dorsal horn were increased significantly in the STZ-H rats (P<0.05), whereas the neuronal number did not change significantly (P>0.05). Parameters of STZ-N rats showed no significant changes. In conclusion, PDN, a form of neuropathic pain, is also associated with a synaptic plasticity (numerical increase) in the spinal dorsal horn. This numerical change might be the reason for central sensitization resulting in reduced pain threshold, enhanced responsiveness, and expanded receptive fields associated with PDN. Therefore, our studies indicate that neuropathic pain conditions with different etiologies might show the same synaptic numerical plasticity changes.

TNF-α Differentially Regulates Synaptic Plasticity in the Hippocampus and Spinal Cord by Microglia-Dependent Mechanisms after Peripheral Nerve Injury

Clinical studies show that chronic pain is accompanied by memory deficits and reduction in hippocampal volume. Experimental studies show that spared nerve injury (SNI) of the sciatic nerve induces long-term potentiation (LTP) at C-fiber synapses in spinal dorsal horn, but impairs LTP in the hippocampus. The opposite changes may contribute to neuropathic pain and memory deficits, respectively. However, the cellular and molecular mechanisms underlying the functional synaptic changes are unclear. Here, we show that the dendrite lengths and spine densities are reduced significantly in hippocampal CA1 pyramidal neurons, but increased in spinal neurokinin-1-positive neurons in mice after SNI, indicating that the excitatory synaptic connectivity is reduced in hippocampus but enhanced in spinal dorsal horn in this neuropathic pain model. Mechanistically, tumor necrosis factor-alpha (TNF-α) is upregulated in bilateral hippocampus and in ipsilateral spinal dorsal horn, whereas brain-derived neurotrophic factor (BDNF) is decreased in the hippocampus but increased in the ipsilateral spinal dorsal horn after SNI. Importantly, the SNI-induced opposite changes in synaptic connectivity and BDNF expression are prevented by genetic deletion of TNF receptor 1 <i>in vivo</i> and are mimicked by TNF-α in cultured slices. Furthermore, SNI activated microglia in both spinal dorsal horn and hippocampus; pharmacological inhibition or genetic ablation of microglia prevented the region-dependent synaptic changes, neuropathic pain, and memory deficits induced by SNI. The data suggest that neuropathic pain involves different structural synaptic alterations in spinal and hippocampal neurons that are mediated by overproduction of TNF-α and microglial activation and may underlie chronic pain and memory deficits. <b>SIGNIFICANCE STATEMENT</b> Chronic pain is often accompanied by memory deficits. Previous studies have shown that peripheral nerve injury produces both neuropathic pain and memory deficits and induces long-term potentiation (LTP) at C-fiber synapses in spinal dorsal horn (SDH) but inhibits LTP in hippocampus. The opposite changes in synaptic plasticity may contribute to chronic pain and memory deficits, respectively. However, the structural and molecular bases of these alterations of synaptic plasticity are unclear. Here, we show that the complexity of excitatory synaptic connectivity and brain-derived neurotrophic factor (BDNF) expression are enhanced in SDH but reduced in the hippocampus in neuropathic pain and the opposite changes depend on tumor necrosis factor-alpha/tumor necrosis factor receptor 1 signaling and microglial activation. The region-dependent synaptic alterations may underlie chronic neuropathic pain and memory deficits induced by peripheral nerve injury.

TNF-α Differentially Regulates Synaptic Plasticity in the Hippocampus and Spinal Cord by Microglia-Dependent Mechanisms after Peripheral Nerve Injury.

Chronic pain is often accompanied by memory deficits. Previous studies have shown that peripheral nerve injury produces both neuropathic pain and memory deficits and induces long-term potentiation (LTP) at C-fiber synapses in spinal dorsal horn (SDH) but inhibits LTP in hippocampus. The opposite changes in synaptic plasticity may contribute to chronic pain and memory deficits, respectively. However, the structural and molecular bases of these alterations of synaptic plasticity are unclear. Here, we show that the complexity of excitatory synaptic connectivity and brain-derived neurotrophic factor (BDNF) expression are enhanced in SDH but reduced in the hippocampus in neuropathic pain and the opposite changes depend on tumor necrosis factor-alpha/tumor necrosis factor receptor 1 signaling and microglial activation. The region-dependent synaptic alterations may underlie chronic neuropathic pain and memory deficits induced by peripheral nerve injury.

Bulleyaconitine A depresses neuropathic pain and potentiation at C-fiber synapses in spinal dorsal horn induced by paclitaxel in rats

Paclitaxel, a widely used chemotherapeutic agent, often induces painful peripheral neuropathy and at present no effective drug is available for treatment of the serious side effect. Here, we tested if intragastrical application of bulleyaconitine A (BLA), which has been approved for clinical treatment of chronic pain in China since 1985, could relieve the paclitaxel-induced neuropathic pain. A single dose of BLA attenuated the mechanical allodynia, thermal hyperalgesia induced by paclitaxel dose-dependently. Repetitive administration of the drug (0.4 and 0.8mg/kg, t.i.d. for 7 d) during or after paclitaxel treatment produced a long-lasting inhibitory effect on thermal hyperalgesia, but not on mechanical allodynia. In consistency with the behavioral results, in vivo electrophysiological experiments revealed that spinal synaptic transmission mediated by C-fiber but not A fiber was potentiated, and the magnitude of long-term potentiation (LTP) at C-fiber synapses induced by the same high frequency stimulation was ~50% higher in paclitaxel-treated rats, compared to the naïve rats. Spinal or intravenous application of BLA depressed the spinal LTP, dose-dependently. Furthermore, patch clamp recordings in spinal cord slices revealed that the frequency but not amplitude of both spontaneous excitatory postsynaptic current (sEPSCs) and miniature excitatory postsynaptic currents (mEPSCs) in lamina II neurons was increased in paclitaxel-treated rats, and the superfusion of BLA reduced the frequency of sEPSCs and mEPSCs in paclitaxel-treated rats but not in naïve ones. Taken together, we provide novel evidence that BLA attenuates paclitaxel-induced neuropathic pain and that depression of spinal LTP at C-fiber synapses via inhibiting presynaptic transmitter release may contribute to the effect.

Long-term potentiation at spinal C-fiber synapses: a target for pathological pain.

Long-term potentiation (LTP), referring to a lasting increase in efficacy of synaptic transmission, is a common mechanism of memory storage in central nervous system (CNS). LTP at C-fiber synapses in spinal dorsal horn is considered as a synaptic model of pathological pain, as the spinal LTP is only induced by noxious electrical and natural stimuli but not by innoxious ones and LTPinducible stimulation is capable of leading to lasting behavioral signs of pathological pain in human and in animals. The molecular mechanisms of spinal LTP at C-fiber synapses are similar to hippocampal LTP in following aspects. Induction of LTP depends on postsynaptic Ca(2+) rise resulting from opening of N-methyl-D-aspartate channels (NMDA) and voltage-gated calcium channels (VGCCs), and Ca(2+) release from intracellular store; Early-phase LTP (<3h) needs activation of intracellular protein kinase A (PKA), protein kinase C (PKC), calcium/calmodulin-dependent protein kinase II (CaMKII), phospholipase C (PLC) and release of nitric oxide (NO); Late-phase LTP (>3h) is dependent on de novo protein synthesis; Activation of either dopamine D1 receptors or PKA, and extrogenous brain-derived neurotrophic factor (BDNF) or ATP directly induces late-phase LTP. Therefore, the drugs targeting at the above molecules may impair memory function of hippocampus. The striking difference between hippocampal LTP and spinal LTP at C-fiber synapses is that activation of glial cells and the over-expression of proinflammatory cytokines, such as tumor necrosis factor-alpha (TNF-α) and interleukin- beta (IL-1β), inhibit LTP in hippocampus, but promote LTP in spinal dorsal horn. The drugs targeting at the neuroinflammatory process may not only attenuate pathological pain but also improve memory in hippocampus.

Ion Channel Modulation in Spinal/Trigeminal Synaptic Plasticity

Ion Channel Modulation in Spinal/Trigeminal Synaptic Plasticity

The direction of synaptic plasticity mediated by C-fibers in spinal dorsal horn is decided by Src-family kinases in microglia: The role of tumor necrosis factor-α

Previous studies have shown that Src-family kinases (SFKs) are selectively activated in spinal microglia following peripheral nerve injury and the activated SFKs play a key role for the development of neuropathic pain. To investigate the underlying mechanism, in the present study the effect of SFKs on long-term potentiation (LTP) at C-fiber synapses in spinal dorsal horn, which is believed as central mechanism of neuropathic pain, was investigated in adult rats. Electrophysiological data revealed that pretreatment with either microglia inhibitor (minocycline, 200μM) or SFKs inhibitors (PP2, 100μM and SU6656, 200μM) reversed the effect of high frequency stimulation (HFS), that is, HFS, which induces long-term potentiation (LTP) normally, induced long-term depression (LTD) after inhibition of either microglia or SFKs. Western blotting analysis showed that the level of phosphorylated SFKs (p-SFKs) in ipsilateral spinal dorsal horn was transiently increased after LTP induced by HFS, starting at 15min and returning to control level at 60min after HFS. Double-labeled immunofluorescence staining demonstrated that p-SFKs were highly restricted to microglia. Furthermore, we found that the inhibitory effects of minocycline or SU6656 on spinal LTP were reversed by spinal application of rat recombinant tumor necrosis factor-α (TNF-α 0.5ng/ml, 200μl). HFS failed to induce LTP of C-fiber evoked field potentials in TNF receptor-1 knockout mice and in rats pretreated with TNF-α neutralization antibody (0.6μg/ml, 200μl). The results suggested that in spinal dorsal horn activation of SFKs in microglia might control the direction of plastic changes at C-fiber synapses and TNF-α might be involved in the process.

Short-Term Plasticity at Primary Afferent Synapse in Rat Spinal Dorsal Horn and Its Biological Function

Short-term plasticity (STP) is an important element of information processing in neuronal networks. As the first synaptic relay between primary afferent fibers (PAFs) and central neurons, primary afferent synapses in spinal dorsal horn (DH) are essential to the initial processing of somatosensory information. In this research, we examined the STP between Aδ-PAFs and spinal DH neurons by patch-clamp recording. Our results showed that depression dominated the STP at primary afferent synapses. The curves of STP had no significant changes in the presence of bicuculline, CTZ or AP-5. Lowering extracellular Ca²⁺ concentration ([Ca²⁺]_o) from 2.4 to 0.8 mM reduced the depression of synaptic responses at all stimulus rates, while raising [Ca²⁺]_o from 2.4 to 4.0 mM increased the synaptic depression. Increasing the bath temperature from 24 to 32°C clearly reduced the depression of all responses. These results indicate that the observed STP is of presynaptic origin and depends on transmitter release. By fitting the experimental data recorded under different conditions, a model of STP was used to quantitatively characterize the observed STP and to analyze the possible mechanisms underlying the effects of [Ca²⁺]_o and temperature. Furthermore, using a model neuron receiving synaptic inputs, we found that with this form of STP, postsynaptic DH neurons could detect rate changes in both rapidly- and slowly-firing afferents with equal sensitivity. The present study links the intrinsic STP properties of primary afferent synapses with their role in processing neural information, and provides a basis for further research on the STP in spinal DH and its biological function under in vivo conditions.