I am a
Home I AM A Search Login

Animal Studies

Share this

Exercise training modulates glutamic acid decarboxylase-65/67 expression through TrkB signaling to ameliorate neuropathic pain in rats with spinal cord injury.

Neuropathic pain is one of the most frequently stated complications after spinal cord injury. In post-spinal cord injury, the decrease of gamma aminobutyric acid synthesis within the distal spinal cord is one of the main causes of neuropathic pain. The predominant research question of this study was whether exercise training may promote the expression of glutamic acid decarboxylase-65 and glutamic acid decarboxylase-67, which are key enzymes of gamma aminobutyric acid synthesis, within the distal spinal cord through tropomyosin-related kinase B signaling, as its synthesis assists to relieve neuropathic pain after spinal cord injury. Animal experiment was conducted, and all rats were allocated into five groups: Sham group, SCI/PBS group, SCI-TT/PBS group, SCI/tropomyosin-related kinase B-IgG group, and SCI-TT/tropomyosin-related kinase B-IgG group, and then T10 contusion SCI model was performed as well as the tropomyosin-related kinase B-IgG was used to block the tropomyosin-related kinase B activation. Mechanical withdrawal thresholds and thermal withdrawal latencies were used for assessing pain-related behaviors. Western blot analysis was used to detect the expression of brain-derived neurotrophic factor, tropomyosin-related kinase B, CREB, p-REB, glutamic acid decarboxylase-65, and glutamic acid decarboxylase-67 within the distal spinal cord. Immunohistochemistry was used to analyze the distribution of CREB, p-CREB, glutamic acid decarboxylase-65, and glutamic acid decarboxylase-67 within the distal spinal cord dorsal horn. The results showed that exercise training could significantly mitigate the mechanical allodynia and thermal hyperalgesia in post-spinal cord injury and increase the synthesis of brain-derived neurotrophic factor, tropomyosin-related kinase B, CREB, p-CREB, glutamic acid decarboxylase-65, and glutamic acid decarboxylase-67 within the distal spinal cord. After the tropomyosin-related kinase B signaling was blocked, the analgesic effect of exercise training was inhibited, and in the SCI-TT/tropomyosin-related kinase B-IgG group, the synthesis of CREB, p-CREB, glutamic acid decarboxylase-65, and glutamic acid decarboxylase-67 within the distal spinal cord were also significantly reduced compared with the SCI-TT/PBS group. This study shows that exercise training may increase the glutamic acid decarboxylase-65 and glutamic acid decarboxylase-67 expression within the spinal cord dorsal horn through the tropomyosin-related kinase B signaling, and this mechanism may play a vital role in relieving the neuropathic pain of rats caused by incomplete SCI.

Learn More >

Low intrinsic efficacy for G protein activation can explain the improved side effect profiles of new opioid agonists.

Biased agonism at G protein-coupled receptors describes the phenomenon whereby some drugs can activate some downstream signaling activities to the relative exclusion of others. Descriptions of biased agonism focusing on the differential engagement of G proteins versus β-arrestins are commonly limited by the small response windows obtained in pathways that are not amplified or are less effectively coupled to receptor engagement, such as β-arrestin recruitment. At the μ-opioid receptor (MOR), G protein-biased ligands have been proposed to induce less constipation and respiratory depressant side effects than opioids commonly used to treat pain. However, it is unclear whether these improved safety profiles are due to a reduction in β-arrestin-mediated signaling or, alternatively, to their low intrinsic efficacy in all signaling pathways. Here, we systematically evaluated the most recent and promising MOR-biased ligands and assessed their pharmacological profile against existing opioid analgesics in assays not confounded by limited signal windows. We found that oliceridine, PZM21, and SR-17018 had low intrinsic efficacy. We also demonstrated a strong correlation between measures of efficacy for receptor activation, G protein coupling, and β-arrestin recruitment for all tested ligands. By measuring the antinociceptive and respiratory depressant effects of these ligands, we showed that the low intrinsic efficacy of opioid ligands can explain an improved side effect profile. Our results suggest a possible alternative mechanism underlying the improved therapeutic windows described for new opioid ligands, which should be taken into account for future descriptions of ligand action at this important therapeutic target.

Learn More >

Bone cancer-induced pain is associated with glutamate signalling in peripheral sensory neurons.

Learn More >

Role of Intraganglionic Transmission in the Trigeminovascular Pathway.

Migraine is triggered by poor air quality and odors through unknown mechanisms. Activation of the trigeminovascular pathway by environmental irritants may occur via activation of TRPA1 receptors on nasal trigeminal neurons, but how that results in peripheral and central sensitization is unclear. The anatomy of the trigeminal ganglion suggests that noxious nasal stimuli are not being transduced to the meninges by axon reflex but likely through intraganglionic transmission. Consistent with this concept, we injected CGRP, ATP or glutamate receptor antagonists or a gap junction channel blocker directly and exclusively into the trigeminal ganglion and blocked meningeal blood flow changes in response to acute nasal TRP agonists. Previously, we observed chronic sensitization of the trigeminovascular pathway after acrolein exposure, a known TRPA1 receptor agonist. To explore the mechanism of this sensitization, we utilized laser dissection microscopy to separately harvest nasal and meningeal trigeminal neuron populations in the absence or presence of acrolein exposure. mRNA levels of neurotransmitters important in migraine were then determined by RT-PCR. TRPA1 message levels were significantly increased in meningeal cell populations following acrolein exposure compared to room air exposure. This was specific to TRPA1 message in meningeal cell populations as changes were not observed in either nasal trigeminal cell populations or dorsal root ganglion populations. Taken together, this data suggests an important role for intraganglionic transmission in acute activation of the trigeminovascular pathway. It also supports a role for upregulation of TRPA1 receptors in peripheral sensitization and a possible mechanism for chronification of migraine after environmental irritant exposure.

Learn More >

Neuroinflammation and central PI3K/Akt/mTOR signal pathway contribute to bone cancer pain.

Pain is one of the most common and distressing symptoms suffered by patients with progression of cancer; however, the mechanisms responsible for hyperalgesia are not well understood. Since the midbrain periaqueductal gray (PAG) is an important component of the descending inhibitory pathway controlling on central pain transmission, in this study we examined the role for pro-inflammatory cytokines (PICs) of the PAG in regulating mechanical and thermal hyperalgesia evoked by bone cancer via PI3K-mTOR signals. Breast sarcocarcinoma Walker 256 cells were implanted into the tibia bone cavity of rats to induce mechanical and thermal hyperalgesia. Western blot analysis and ELISA were used to examine PI3K/Akt/mTOR and PIC receptors and the levels of IL-1β, IL-6 and TNF-α. Protein expression levels of p-PI3K/p-Akt/p-mTOR were amplified in the PAG of bone cancer rats; and blocking PI3K-mTOR pathways in the PAG attenuated hyperalgesia responses. In addition, IL-1β, IL-6 and TNF-α were elevated in the PAG of bone cancer rats and expression of their respective receptors (namely, IL-1R, IL-6R and TNFR subtype TNFR1) was upregulated. Inhibition of IL-1R, IL-6R and TNFR1 alleviated mechanical and thermal hyperalgesia in bone cancer rats, accompanied with downregulated PI3K-mTOR. Our data suggest that upregulation of PIC signal in the PAG of cancer rats amplifies PI3K-mTOR signal in this brain region and alters the descending pathways in regulating pain transmission; and this thereby contributes to the development of bone cancer-induced pain.

Learn More >

Temporal and sex differences in the role of BDNF/TrkB signaling in hyperalgesic priming in mice and rats.

Brain-derived neurotrophic factor (BDNF) signaling through its cognate receptor, TrkB, is a well-known promoter of synaptic plasticity at nociceptive synapses in the dorsal horn of the spinal cord. Existing evidence suggests that BDNF/TrkB signaling in neuropathic pain is sex dependent. We tested the hypothesis that the effects of BDNF/TrkB signaling in hyperalgesic priming might also be sexually dimorphic. Using the incision postsurgical pain model in male mice, we show that BDNF sequestration with TrkB-Fc administered at the time of surgery blocks the initiation and maintenance of hyperalgesic priming. However, when BDNF signaling was blocked prior to the precipitation of hyperalgesic priming with prostaglandin E (PGE), priming was not reversed. This result is in contrast to our findings in male mice with interleukin-6 (IL6) as the priming stimulus where TrkB-Fc was effective in reversing the maintenance of hyperalgesic priming. Furthermore, in IL6-induced hyperalgesic priming, the BDNF sequestering agent, TrkB-fc, was effective in reversing the maintenance of hyperalgesic priming in male mice; however, when this experiment was conducted in female mice, we did not observe any effect of TrkB-fc. This markedly sexual dimorphic effect in mice is consistent with recent studies showing a similar effect in neuropathic pain models. We tested whether the sexual dimorphic role for BDNF was consistent across species. Importantly, we find that this sexual dimorphism does not occur in rats where TrkB-fc reverses hyperalgesic priming fully in both sexes. Finally, to determine the source of BDNF in hyperalgesic priming in mice, we used transgenic mice (  ×  mice) with BDNF eliminated from microglia. From these experiments we conclude that BDNF from microglia does not contribute to hyperalgesic priming and that the key source of BDNF for hyperalgesic priming is likely nociceptors in the dorsal root ganglion. These experiments demonstrate the importance of testing mechanistic hypotheses in both sexes in multiple species to gain insight into complex biology underlying chronic pain.

Learn More >

Roles of TNF-α and IL-6 in regulating bone cancer pain via TRPA1 signal pathway and beneficial effects of inhibition of neuro-inflammation and TRPA1.

Pain is one of the most common and distressing symptoms suffered by patients with progression of bone cancer; however, the mechanisms responsible for hyperalgesia are not well understood. The purpose of our current study was to determine contributions of the sensory signaling pathways of inflammatory tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6), and downstream transient receptor potential ankyrin 1 (TRPA1) to neuropathic pain induced by bone cancer. We further determined if influencing these pathways can improve bone cancer pain. Breast sarcocarcinoma Walker 256 cells were implanted into the tibia bone cavity of rats to induce mechanical and thermal hyperalgesia. ELISA and western blot analysis were used to examine 1) the levels of TNF-α and IL-6 in dorsal root ganglion (DRG); and 2) protein expression of TNF-α and IL-6 receptors (TNFR1 and IL-6R) and TRPA1 as well as intracellular signals (p38-MAPK and JNK). TNF-α and IL-6 were elevated in the DRG of bone cancer rats and expression of TNFR1, IL-6R and TRPA1 was upregulated. In addition, inhibition of TNFR1 and IL-6R alleviated mechanical and thermal hyperalgesia in bone cancer rats, accompanied with downregulated TRPA1 and p38-MAPK and JNK. We revealed specific signaling pathways leading to neuropathic pain during the development of bone cancer, including TNF-α-TRPA1 and IL-6-TRPA1 signal pathways. Overall, our data suggest that blocking these signals is beneficial to alleviate bone cancer pain.

Learn More >

The Effects of Acute Neonatal Pain on Expression of Corticotropin Releasing Hormone and Juvenile Anxiety in a Rodent Model.

Premature infants in the Neonatal Intensive Care Unit (NICU) may be subjected to numerous painful procedures without analgesics. One necessary, though acutely painful, procedure is the use of heel lances to monitor blood composition. The current study examined the acute effects of neonatal pain on maternal behavior as well as amygdalar and hypothalamic activation, and the long-term effects of neonatal pain on later-life anxiety-like behavior, using a rodent model. Neonatal manipulations consisted of either painful needle pricks or non-painful tactile stimulation in subjects' left plantar paw surface which occurred four times daily during the first week of life (PND 1 – 7). Additionally, maternal behaviors in manipulated litters were compared against undisturbed litters via scoring of videotaped interactions to examine the long-term effects of pain on dam-pup interactions. Select subjects underwent neonatal brain collection (PND 6) and fluorescent hybridization (FISH) for corticotropin releasing hormone (CRH) and the immediate early gene c-fos. Other subjects were raised to juvenile age (PND 24 and PND 25) and underwent innate anxiety testing utilizing an elevated plus maze protocol. FISH indicated that neonatal pain influenced amygdalar CRH and c-fos expression, predominately in males. No significant increase in c-fos or CRH expression was observed in the hypothalamus. Additionally, neonatal pain altered anxiety behaviors independent of sex, with neonatal pain subjects showing the highest frequency of exploratory behavior. Neonatal manipulations did not alter maternal behaviors. Overall, neonatal pain drives CRH expression and produces behavioral changes in anxiety that persist until the juvenile stage. This report expands on current rodent model research performed to assess the long-term effects of highly utilized neonatal intensive care unit (NICU) procedures. The NICU plays an integral role in pediatric medicine by significantly reducing infant mortality and providing necessary procedures to preterm or unwell newborns. However, procedures in the NICU are often stressful and painful. A common procedure performed in the NICU is heel lances to monitor blood composition. This, along with numerous other painful procedures, are often performed on NICU babies without the benefit of analgesics. Our study identifies key neurological indicators which are altered in response to neonatal pain. Additionally, we explore the later anxiety of subjects exposed to neonatal pain.

Learn More >

Analgesic effects of optogenetic inhibition of basolateral amygdala inputs into the prefrontal cortex in nerve injured female mice.

Peripheral nerve injury can lead to remodeling of brain circuits, and this can cause chronification of pain. We have recently reported that male mice subjected to spared injury of the sciatic nerve undergo changes in the function of the medial prefrontal cortex (mPFC) that culminate in reduced output of layer 5 pyramidal cells. More recently, we have shown that this is mediated by alterations in synaptic inputs from the basolateral amygdala (BLA) into GABAergic interneurons in the mPFC. Optogenetic inhibition of these inputs reversed mechanical allodynia and thermal hyperalgesia in male mice. It is known that the processing of pain signals can exhibit marked sex differences. We therefore tested whether the dysregulation of BLA to mPFC signaling is equally altered in female mice. Injection of AAV-Arch3.0 constructs into the BLA followed by implantation of a fiberoptic cannula into the mPFC in sham and SNI operated female mice was carried out, and pain behavioral responses were measured in response to yellow light mediated activation of this inhibitory opsin. Our data reveal that Arch3.0 activation leads to a marked increase in paw withdrawal thresholds and latencies in response to mechanical and thermal stimuli, respectively. However, we did not observe nerve injury-induced changes in mPFC layer 5 pyramidal cell output in female mice. Hence, the observed light-induced analgesic effects may be due to compensation for dysregulated neuronal circuits downstream of the mPFC.

Learn More >

IFNβ treatment inhibits nerve injury-induced mechanical allodynia and MAPK signaling by activating ISG15 in mouse spinal cord.

Neuropathic pain is difficult to treat and remains a major clinical challenge worldwide. While the mechanisms which underlie the development of neuropathic pain are incompletely understood, interferon signaling by the immune system is known to play a role. Here, we demonstrate a role for IFNβ in attenuating mechanical allodynia induced by the spared nerve injury in mice. The results show that intrathecal administration of IFNβ (dosages up to 5000U) produces significant, transient, and dose-dependent attenuation of mechanical allodynia without observable effects on motor activity or feeding behavior, as is common with IFN administration. This analgesic effect is mediated by the ubiquitin-like protein ISG15, which is potently induced within the spinal cord following intrathecal delivery of IFNβ. Both free and conjugated ISG15 are elevated following IFNβ treatment, and this effect is increased in UBP43 mice lacking a key deconjugating enzyme. The IFNβ-mediated analgesia reduces MAPK signaling activation following nerve injury, and this effect requires induction of ISG15. These findings highlight a new role for IFNβ, ISG15 and MAPK signaling in immunomodulation of neuropathic pain and may lead to new therapeutic possibilities. Perspective: Neuropathic pain is frequently intractable in a clinical setting, and new treatment options are needed. Characterizing the anti-nociceptive potential of IFNβ and the associated downstream signaling pathways in preclinical models may lead to the development of new therapeutic options for debilitating neuropathies.

Learn More >

Search