. An automated flinch detecting system for use in the formalin nociceptive bioassay. J Appl Physiol 90: [2386][2387][2388][2389][2390][2391][2392][2393][2394][2395][2396][2397][2398][2399][2400][2401][2402] 2001.-The biphasic display of pawflinch behavior in the rat after injection of formalin into the dorsum of the hind paw is used for the screening of antihyperalgesic agents. Described and characterized here is a less labor-intensive system for counting flinch activity by detecting movement of a small metal band placed on the formalin-injected paw. A signal is generated as the band breaks the electromagnetic field of a loop antenna located under the rat and processed through an algorithm that determines flinch activity using 1) amplitude, 2) zero-voltage crossing, and 3) signal duration. Flinches are summed and stored over a selected collection interval throughout the assay for later analysis. Studies have validated the measures with respect to 1) system stability over time; 2) system-to-"practiced observer" correlation on flinch detection, r 2 ϭ 0.94; 3) system variables including time of day, sex, age, and body weight; and 4) 50% effective dose values similar to those previously reported for intrathecal morphine and the NMDA antagonist MK-801. formalin test; spinal sensitization; pain models; flinching behavior THE ESCAPE RESPONSE OR AGITATION evoked by a transient, strong stimulus attests to there being a close relationship between stimulus intensity, peripheral afferent discharge, and magnitude of the pain state as defined by response latency and magnitude. There are situations, however, in which the magnitude of the response to pain may exceed what would normally be anticipated, given the magnitude of the physical stimulus and the afferent traffic generated by that stimulus (31,45,47). These situations are loosely considered as reflecting a state of hyperalgesia, possibly arising from sensitization of the peripheral terminal and/or a central facilitation.Several preclinical models have been developed that may reflect the significance played by such facilitation on behavior. The common characteristic found in these models is the injury that is induced and its causing of the sensory axon to produce a persistent discharge. A frequently used method of producing injury in the rat is the subcutaneous injection of a small volume of irritant such as formalin into its hind paw. Typically, after the formalin injection, the rat displays a biphasic (phase I and phase II) incidence of flinching (rapid paw shaking) and licking of the injected paw (18,42,43). The behavioral syndrome produced by the injection of formalin into the paw has been widely used to define the pharmacology of systems that regulate facilitated processing. The "formalin test" has evolved into a widely used tool in the screening of analgesic and anti-hyperalgesic drugs (45).An important limitation of this behavioral model is its labor-intensive nature regarding data collection and the time required to train observers in its reliable implementation. ...

Xen2174 is a structural analogue of Mr1A, a chi-conopeptide recently isolated from the venom of the marine cone snail, Conus marmoreus. Although both chi-conopeptides are highly selective inhibitors of the norepinephrine transporter (NET), Xen2174 has superior chemical stability relative to Mr1A. It is well-known that tricyclic antidepressants (TCAs) are also potent NET inhibitors, but their poor selectivity relative to other monoamine transporters and various G-protein-coupled receptors, results in dose-limiting side-effects in vivo. As TCAs and the alpha(2)-adrenoceptor agonist, clonidine, have established efficacy for the relief of neuropathic pain, this study examined whether intrathecal (i.t.) Xen2174 alleviated mechanical allodynia in rats with either a chronic constriction injury of the sciatic nerve (CCI-rats) or an L5/L6 spinal-nerve injury. The anti-allodynic responses of i.t. Mr1A and i.t. morphine were also investigated in CCI-rats. Paw withdrawal thresholds were assessed using calibrated von Frey filaments. Bolus doses of i.t. Xen2174 produced dose-dependent relief of mechanical allodynia in CCI-rats and in spinal nerve-ligated rats. Dose-dependent anti-allodynic effects were also produced by i.t. bolus doses of Mr1A and morphine in CCI-rats, but a pronounced 'ceiling' effect was observed for i.t. morphine. The side-effect profiles were mild for both chi-conopeptides with an absence of sedation. Confirming the noradrenergic mechanism of action, i.t. co-administration of yohimbine (100 nmol) with Xen2174 (10 nmol) abolished Xen2174s anti-allodynic actions. Xen2174 appears to be a promising candidate for development as a novel therapeutic for i.t. administration to patients with persistent neuropathic pain.

Contulakin-G is a novel conopeptide with an incompletely defined mechanism of action. To assess nociceptive activity we delivered Contulakin-G as a bolus intrathecally (0.03, 0.1, 0.3, 3 nmol) or epidurally (10, 30, 89 nmol) in rats. Intrathecal Contulakin G significantly decreased Phase II and, to a lesser degree, Phase I paw flinching produced by intradermal formalin. Intrathecal and epidural doses of ED50s were 0.07 nmol and 45 nmol, respectively, giving an epidural/intrathecal ED50 ratio = 647). In dogs, intrathecal Contulakin-G (50-500 nmoL) produced a dose-dependent increase in the thermally evoked skin twitch latency by 30 min after administration, as did morphine (150 and 450 nmol). Epidural morphine (750 and 7500 nmol), but not epidural 1000 nmol Contulakin-G, also significantly decreased skin twitch in dogs. No changes in motor function were seen in any rats or dogs receiving these doses of Contulakin-G. In dogs, no physiologically significant dose-dependent changes in motor function, heart rate, arterial blood pressure, or body temperature were found. Contulakin-G is a potent antinociceptive drug when delivered intrathecally with no observable negative side effects in rats or dogs and may provide an alternative to opioid spinal analgesics.