Calcium influx through neuronal voltage-sensitive calcium channels (VSCC S) mediates nociceptive information in the spinal dorsal horn. In fact, spinally administered VSCC S blockers, such as omega-conotoxin MVIIA, have analgesic effect apart of their low therapeutic index and many side effects. Here we study the analgesic potential of Ph alpha 1beta, a calcium channel blocker, in rodent models of acute and persistent pain. Spinally administered Ph alpha 1beta showed higher efficacy and long-lasting analgesia in a thermal model of pain, when compared with omega-conotoxin MVIIA. Moreover, Ph alpha 1beta was more effective and potent than omega-conotoxin MVIIA not only to prevent, but especially to reverse, previously installed persistent chemical and neuropathic pain. Furthermore, the analgesic action of both toxins are related with the inhibition of Ca2+-evoked release of pro-nociceptive neurotransmitter, glutamate, from rat spinal cord synaptosomes and decrease of glutamate overflow in cerebrospinal fluid. When side effects were assessed, we found that Ph alpha 1beta had a therapeutic index wider than omega- conotoxin MVIIA. Finally, recombinant Ph alpha 1beta expressed in Escherichia coli showed marked analgesic activity similar to the native toxin. Taken together, the present study demonstrates that native and recombinant Ph alpha 1beta have analgesic effects in rodent models of pain, suggesting that this toxin may have potential to be used as a drug in the control of persistent pathological pain.

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Four neurotoxic polypptidcs were purified from the venom of the South American 'armed' spider Phuncmiu nigrivenrer (Keys) by gel filtration and rcvcrse phase FPLC and HPLC. These cystcine-rich polypcptidcs exhibited dimerent lcvcls of neurotoxicity in mice after intraccrebrovcntricular injection. Tic'-I, TxZ5 and Tx2-6 caused spastic paralysis nnd deuh, but the less toxic Tx2-9 produced only tail erection and scratching. The molecular weights of the polypeptides as determined by desorption mass spectroscoopy were 5838.8 for Tx2-1, 5116.6 (Txl-5). 5291.3 (Tx2-6) and 3742.1 (Tx2-9). The complele amino acid sequences of the neurotoxins were determhxd by automnted Edman degradation and by manual DABITC-PITC microscqeuncc analysis of peptidcs obtained atIer digestions with various proteases. The amino acid sequences ofTx2.1 (53 residue), Tx2-5 (49 residues) and Tx2-6 (48 residues) were homologous, but had only limited similarities to the less toxic Tx2-9 (32 residues). All four polypeptides bad varying seqlxnce identities with other neurotoxins from different spider species and biologiiwlly active peptides from scorpions, a sea snail and seeds of Mirrrliitijufupa.Spider venom; Amino acid xquence; Neuroloxin; Phoneutriu rri~rivwkr

The treatment with the chemotherapeutic agent paclitaxel produces a painful peripheral neuropathy, and is associated with an acute pain syndrome in a clinically significant number of patients. However, no standard therapy has been established to manage the acute pain or the chronic neuropathic pain related to paclitaxel. In the present study, we evaluated the analgesic potential of two N-type voltage-gated calcium channel (VGCC) blockers, ω-conotoxin MVIIA and Phα1β, on acute and chronic pain induced by paclitaxel. Adult male rats were treated with four intraperitoneal injections of paclitaxel (1+1+1+1mg/kg, in alternate days) and the development of mechanical hyperalgesia was evaluated 24h (acute painful stage) or 15days (chronic painful stage) after the first paclitaxel injection. Not all animals showed mechanical hyperalgesia 24h after the first paclitaxel injection, but those that showed developed a more intense mechanical hyperalgesia at the chronic painful stage. Intrathecal administration (i.t.) of ω-conotoxin MVIIA (3-300pmol/site) or Phα1β (10-300pmol/site) reduced the mechanical hyperalgesia either at the acute or at the chronic painful stage induced by paclitaxel. When administered at the acute painful stage, ω-conotoxin MVIIA (300pmol/site, i.t.) and Phα1β (300pmol/site, i.t.) prevented the worsening of chronic mechanical hyperalgesia. Furthermore, Phα1β (30-300pmol/site, i.t.) elicited less adverse effects than ω-conotoxin MVIIA (10-300 pmol/site, i.t.). Taken together, our data evidence the involvement of N-type VGCC in pain sensitization induced by paclitaxel and point out the potential of Phα1β as a safer alternative than ω-conotoxin MVIIA to treat the pain related to paclitaxel.

The marine snail peptide ziconotide (x-conotoxin MVIIA) is used as an analgesic in cancer patients refractory to opioids, but may induce severe adverse effects. Animal venoms represent a rich source of novel drugs, so we investigated the analgesic effects and the side-effects of spider peptide Pha1b in a model of cancer pain in mice with or without tolerance to morphine analgesia. Cancer pain was induced by the inoculation of melanoma B16-F10 cells into the hind paw of C57BL ⁄ 6 mice. After 14 days, painful hypersensitivity was detected and Pha1b or x-conotoxin MVIIA (10-100 pmol ⁄ site) was intrathecally injected to evaluate the development of antinociception and side-effects in control and morphine-tolerant mice. The treatment with Pha1b or x-conotoxin MVIIA fully reversed cancer-related painful hypersensitivity, with long-lasting results, at effective doses 50% of 48 (32-72) or 33 (21-53) pmol ⁄ site, respectively. Pha1b produced only mild adverse effects, whereas x-conotoxin MVIIA induced dose-related side-effects in mice at analgesic doses (estimated toxic dose 50% of 30 pmol ⁄ site). In addition, we observed that Pha1b was capable of controlling cancer-related pain even in mice tolerant to morphine antinociception (100% of inhibition) and was able to partially restore morphine analgesia in such animals (56 AE 5% of inhibition). In this study, Pha1b was as efficacious as x-conotoxin MVIIA in inducing analgesia in a model of cancer pain without producing severe adverse effects or losing efficacy in opioid-tolerant mice, indicating that Pha1b has a good profile for the treatment of cancer pain in patients. (Cancer Sci 2013; 104: 1226-1230