Antonina Stefania Mangione

Effects of CGRP receptor antagonism in nitroglycerin-induced hyperalgesia

Background The release of calcitonin gene-related peptide (CGRP) from trigeminal nerves plays a central role in the pathophysiology of migraine and clinical evidence shows an antimigraine effect for CGRP receptor antagonists. Systemic administration of nitroglycerin (NTG), a nitrovasodilator, consistently provokes spontaneous-like migraine attacks in migraine sufferers; in the rat, systemic NTG induces a condition of hyperalgesia, probably through the activation of cerebral/spinal structures involved in nociceptive transmission. Aim The aim of this article is to test the analgesic effect of the CGRP receptor antagonist MK-8825 in two animal models of pain that may be relevant for migraine: the tail flick test and the formalin test performed during NTG-induced hyperalgesia. Results MK-8825 showed analgesic activity when administered alone at both the tail flick test and the formalin test. Furthermore, the CGRP antagonist proved effective in counteracting NTG-induced hyperalgesia in both tests. MK-8825 indeed reduced the nociceptive behavior when administered either simultaneously or prior to (30–60 minutes before) NTG. Conclusion These data suggest that MK-8825 may represent a potential therapeutic tool for the treatment of migraine.

Effects of peripheral FAAH blockade on NTG-induced hyperalgesia - Evaluation of URB937 in an animal model of migraine

Background Systemic nitroglycerin (NTG) activates brain nuclei involved in nociceptive transmission as well as in neuroendocrine and autonomic functions in rats. These changes are considered relevant for migraine because NTG consistently provokes spontaneous-like migraine attacks in migraineurs. Several studies have suggested a relationship between the endocannabinoid levels and pain mediation in migraine. URB937, a peripheral inhibitor of fatty acid amide hydrolase (FAAH)—the enzyme that degrades anandamide, produces analgesia in animal models of pain, but there is no information on its effects in migraine. Aim We evaluated whether URB937 alters nociceptive responses in the animal model of migraine based on NTG administration in male rats, using the tail flick test and the plantar and orofacial formalin tests, under baseline conditions and after NTG administration. Furthermore, we investigated whether URB937 affects NTG-induced c-Fos expression in the brain. Results During the tail flick test, URB937 showed an antinociceptive effect in baseline conditions and it blocked NTG-induced hyperalgesia. URB937 also proved effective in counteracting NTG-induced hyperalgesia during both the plantar and orofacial formalin tests. Mapping of brain nuclei activated by NTG indicates that URB937 significantly reduces c-Fos expression in the nucleus trigeminalis caudalis and the locus coeruleus. Conclusions The data suggest that URB937 is capable of changing, probably via indirect mechanisms, the functional status of central structures that are important for pain transmission in an animal model of migraine.

IkappaB-alpha expression following transient focal cerebral ischemia is modulated by nitric oxide.

The role of nitric oxide (NO) in cerebral ischemia/reperfusion (IR) has been intensively investigated. In general NO is regarded as a mediator of ischemia-associated neuronal damage, as inhibitors of NO synthesis ameliorate neuronal injury during permanent focal cerebral ischemia, however the exact role of NO in ischemia remains controversial. It has been previously shown that NO-donors can directly inhibit the DNA binding activity of NF-kappaB family proteins and strong evidence supports that activation of NF-κB contributes to ischemia-induced neuronal injury. In this study, we have investigated whether NO production by nNOS, eNOS and iNOS modulates IkB-alpha expression in cerebral ischemia, by using various inhibitors of NOS, in rats subjected to transient (1h) middle cerebral artery occlusion (tMCAo). Male Wistar rats were treated intraperitoneally (i.p.) with 3mg/kg of NG-nitro-l-arginine methyl ester (l-NAME, a non-selective NOS inhibitor), 5mg/kg of l-N6-(1-iminoethyl)-lysine (l-NIL, an inducible NOS inhibitor), 25mg/kg of 7-Nitroindazole (7-NI, a neuronal NOS inhibitor) and 10mg/Kg of l-N-(1-iminoethyl)ornithine (l-NIO, a selective eNOS inhibitor) 15min before the induction of tMCAo. Cortical IkB-alpha expression was evaluated by western blotting and its decrease was considered as an indicator of NF-κB activation. IkB-alpha expression decreased in ischemic cortices when compared with the cortices of the sham group, thus confirming the activation of NF-κB in ischemic conditions. Pre-treatment with l-NAME, l-NIL and 7-NI significantly reduced the infarct volume and prevented ischemia-induced reduction in IkB-alpha expression. Conversely, pretreatment with l-NIO was associated with a significant increase in infarct volume and a reduction in IkB-alpha expression. These findings suggest that NO of neuronal and inducible origin promotes NF-κB activation via IkB-alpha modulation and mediates ischemic-related damage in the brain following ischemia.

Intracarotid Infusion of Mesenchymal Stem Cells in an Animal Model of Parkinson’s Disease, Focusing on Cell Distribution and Neuroprotective and Behavioral Effects

Mesenchymal stem cells (MSCs) have been proposed as a potential therapeutic tool for Parkinsons disease (PD) and systemic administration of these cells has been tested in preclinical and clinical studies. However, no information on survival and actual capacity of MSCs to reach the brain has been provided. In this study, we evaluated homing of intraarterially infused rat MSCs (rMSCs) in the brain of rats bearing a 6‐hydroxydopamine (6‐OHDA)‐induced lesion of the nigrostriatal tract, to establish whether the toxin‐induced damage is sufficient to grant MSC passage across the blood‐brain barrier (BBB) or if a transient BBB disruption is necessary. The rMSC distribution in peripheral organs and the effects of cell infusion on neurodegenerative process and motor deficits were also investigated. rMSCs were infused 14 days after 6‐OHDA injection. A hyperosmolar solution of mannitol was used to transiently permeabilize the BBB. Behavioral impairment was assessed by adjusting step test and response to apomorphine. Animals were sacrificed 7 and 28 days after cell infusion. Our work shows that appreciable delivery of rMSCs to the brain of 6‐OHDA‐lesioned animals can be obtained only after mannitol pretreatment. A notable percentage of infused cells accumulated in peripheral organs. Infusion of rMSCs did not modify the progression of 6‐OHDA‐induced damage or the motor impairment at the stepping test, but induced progressive normalization of the pathological response (contralateral turning) to apomorphine administration. These findings suggest that many aspects should be further investigated before considering any translation of MSC systemic administration into the clinical setting for PD treatment.

Temporal profile of vascular changes induced by systemic nitroglycerin in the meningeal and cortical districts

Background: Clinical studies indicated that nitric oxide (NO) donors cause regional changes in cerebral blood flow (CBF), similar to those reported in spontaneous migraine. Systemic nitroglycerin (NTG), a NO donor, is a well-accepted experimental model of migraine. In this study we have examined the effects of NTG on the meningeal and cortical blood flow in rats. Methods: Regional blood flow was monitored in male Sprague-Dawley rats using laser Doppler flowmetry before and after NTG/saline injection over 150 minutes. The effect of pre-treatment with Nω-nitro-L-arginine ester (L-NAME) or 7-nitroindazole (7-NI) on NTG-induced changes on blood flow was also investigated. Results: In the dura NTG caused a biphasic response represented by an initial decrease in blood flow followed by a significant increase. At variance, in the cortex NTG caused only an increase in blood flow. Pre-treatment with either L-NAME or 7-NI prevented NTG-induced increase in blood flow in both districts, while only L-NAME also prevented NTG-induced decrease in dural blood flow. Conclusion: The present findings provide additional information on the timing of effects of NTG on blood flow at both the meningeal and cortical levels. These effects seem to be related to vasoregulatory mechanisms and/or metabolic activity in response to the synthesis of endogenous NO.

Nonparalytic botulinum molecules for the control of pain.

Abstract Local injections of botulinum toxins have been reported to be useful not only for the treatment of peripheral neuropathic pain and migraine but also to cause long-lasting muscle paralysis, a potentially serious side effect. Recently, a botulinum A-based molecule (“BiTox”) has been synthesized that retains neuronal silencing capacity without triggering muscle paralysis. In this study, we examined whether BiTox delivered peripherally was able to reduce or prevent the increased nociceptive sensitivity found in animal models of inflammatory, surgical, and neuropathic pain. Plasma extravasation and edema were also measured as well as keratinocyte proliferation. No motor deficits were seen and acute thermal and mechanical nociceptive thresholds were unimpaired by BiTox injections. We found reduced plasma extravasation and inflammatory edema as well as lower levels of keratinocyte proliferation in cutaneous tissue after local BiTox injection. However, we found no evidence that BiTox was transported to the dorsal root ganglia or dorsal horn and no deficits in formalin-elicited behaviors or capsaicin or formalin-induced c-Fos expression within the dorsal horn. In contrast, Bitox treatment strongly reduced A-nociceptor-mediated secondary mechanical hyperalgesia associated with either complete Freund’s adjuvant (CFA)-induced joint inflammation or capsaicin injection and the hypersensitivity associated with spared nerve injury. These results imply that although local release of neuromodulators from C-fibers was inhibited by BiTox injection, C-nociceptive signaling function was not impaired. Taken together with recent clinical data the results suggest that BiTox should be considered for treatment of pain conditions in which A-nociceptors are thought to play a significant role.

Selective neuronal silencing using synthetic botulinum molecules alleviates chronic pain in mice

Silencing key neurons with botulinum toxin conjugates exerts long-lasting pain relief in mouse models of chronic pain. Relieving pain with botox Chronic pain affects more than 25 million Americans and is associated with reduced life span, anxiety, and depression. Opioid administration is often effective in relieving pain but, unfortunately, opioids have serious side effects, including risk of addiction and overdose. In a new study, Maiarù et al. have leveraged the inhibitory effects of botulinum toxin on neuronal activity. They developed two botulinum-conjugated molecules (SP-BOT and Derm-BOT) that were able to silence subpopulations of pain-related spinal neurons in several mouse models of chronic pain. Intrathecal administration of one dose of SP-BOT or Derm-BOT produced long-term pain relief in the mouse models that was comparable to the effects of opioid treatment. The results suggest that botulinum-conjugated molecules could be an opioid-free alternative for treating chronic pain. Chronic pain is a widespread debilitating condition affecting millions of people worldwide. Although several pharmacological treatments for relieving chronic pain have been developed, they require frequent chronic administration and are often associated with severe adverse events, including overdose and addiction. Persistent increased sensitization of neuronal subpopulations of the peripheral and central nervous system has been recognized as a central mechanism mediating chronic pain, suggesting that inhibition of specific neuronal subpopulations might produce antinociceptive effects. We leveraged the neurotoxic properties of the botulinum toxin to specifically silence key pain-processing neurons in the spinal cords of mice. We show that a single intrathecal injection of botulinum toxin conjugates produced long-lasting pain relief in mouse models of inflammatory and neuropathic pain without toxic side effects. Our results suggest that this strategy might be a safe and effective approach for relieving chronic pain while avoiding the adverse events associated with repeated chronic drug administration.

P007. Inhibition of monoacylglycerol lipase activity modulates the activation of brain structures relevant for migraine pathogenesis

Author(s): Greco, Rosaria; Bandiera, Tiziano; Mangione, Antonina S; Demartini, Chiara; Nappi, Giuseppe; Sandrini, Giorgio; Piomelli, Daniele; Tassorelli, Cristina

EHMTI-0141. URB937 as a potential therapeutic option for migraine: evaluation in animal model of migraine

Systemic nitroglycerin (NTG) activates cerebral nuclei of rat involved in nociceptive transmission, as well as in neuroendocrine and autonomic functions. These changes are considered relevant for migraine pain, since NTG consistently provokes spontaneous-like migraine attacks in migraineurs. Several reports have suggested the existence of relations between the endocannabinoids and migraine. URB937, a peripheral fatty acid amide hydrolase (FAAH) inhibitor, induces analgesia in animal models of pain but there is no information on its effects in migraine.