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Activation of the KOR produces antipruritic effects. In this study, we investigated three KOR ligands for their abilities to cause scratching behaviors in mice, an animal model of itch, and the involvement of the KOR in these behaviors. Two regioisomers are especially interesting: 6′-guanidinonaltrindole (6′-GNTI, a reportedly KOR G protein-biased agonist) and 5′-guanidinonaltrindole (5′-GNTI, a KOR antagonist). 5′-GNTI is known to precipitate frenzied, excessive scratching in mice. Here we report for the first time that 6′-GNTI causes comparable itching-like behavior in this species, which was not affected by KOR deletion in mice, indicating that it is not mediated by the KOR. We further studied the KOR antagonist LY2456302 (Aticaprant), which is in clinical trials for treatment of substance abuse, for scratching effect in both wild type and KOR knock out mice. Similar numbers of scratches were observed in wild type and KOR knock out mice. We call attention to pruritus as a possible negative side effect of KOR antagonists and 6′-GNTI in humans.

The aims of the present study were to establish if nalfurafine, a kappa opioid agonist, inhibits compulsive scratching in mice elicited by the s.c. administration (behind the neck) of 5′-guanidinonaltrindole (GNTI), a kappa opioid antagonist; to assess if nalfurafine prevents c-fos expression provoked by GNTI or compound 48/80, two chemically diverse pruritogens; and to distinguish on the basis of neuroanatomy, those neurons in the brainstem activated by either GNTI-induced itch or formalin-induced pain (both compounds given s.c. to the right cheek). Pretreatment of mice with nalfurafine (0.001–0.03 mg/kg s.c.) attenuated GNTI (0.3 mg/kg)-evoked scratching dose-dependently. A standard antiscratch dose of nalfurafine (0.02 mg/kg) had no marked effect on the spontaneous locomotion of mice. Tolerance did not develop to the antiscratch activity of nalfurafine. Both GNTI and compound 48/80 provoked c-fos expression on the lateral side of the superficial layer of the dorsal horn of the cervical spinal cord and pretreating mice with nalfurafine inhibited c-fos expression induced by both pruritogens. In contrast to formalin, GNTI did not induce c-fos expression in the trigeminal nucleus suggesting that pain and itch sensations are projected differently along the sensory trigeminal pathway. Our data indicate that the kappa opioid system is involved, at least in part, in the pathogenesis of itch; and that nalfurafine attenuates excessive scratching and prevents scratch-induced neuronal activity at the spinal level. On the basis of our results, nalfurafine holds promise as a potentially useful antipruritic in human conditions involving itch.

The aim of this study was to establish the effect of lidocaine, a local anesthetic, on pain and itch using formalin-induced nociception and kappa opioid antagonist-induced scratching models in mice. We investigated if local intradermal pretreatment (at -10 min) with lidocaine N-ethyl bromide (lidocaine, 2%, 0.1 ml) antagonizes behavioral responses and prevents c-fos expression induced by pain and itch. Male, Swiss Webster mice (25-30 g, n=6-10) were used. Formalin (5%, 20 microl, s.c.) or saline was administered to the right dorsal hindpaw and the time spent licking this paw was recorded at 0-10 min and 20-35 min. For itching, mice were challenged with 5'-guanidinonaltrindole (GNTI, 0.3mg/kg, s.c., behind the neck) or saline and the number of neck-directed scratches with hindpaws was counted for 30 min. C-fos immunohistochemistry was performed in lumbar (for pain) and cervical (for scratching) spinal sections 2h after the respective treatments. We found that lidocaine (a) antagonizes both formalin-induced pain and GNTI-induced scratching and (b) prevents c-fos expression evoked by pain (medial side of the superficial layer and deeper layers of the dorsal horn) and itch (lateral side of the superficial layer of the dorsal horn). Additionally, GNTI caused c-fos activation in mice wearing an Elizabethan collar (to prevent scratching of the neck) suggesting that GNTI provokes c-fos expression by inducing an itch sensation. Our results highlight the antipruritic properties of lidocaine and argue for its comprehensive clinical testing against pruritic states.

In an effort to find novel agents which selectively target the kappa opioid receptor (KOPR), we modified the furan ring of the highly potent and selective KOPR agonist salvinorin A. Introduction of small substituents at C-16 was well tolerated. 12-epi-Salvinorin A, synthesized in four steps from salvinorin A, was a selective partial agonist at the KOPR. No clear SAR patterns were observed for C-13 aryl ketones. Introducing a hydroxymethylene group between C-12 and the furan ring was tolerated. Small C-13 esters and ethers gave weak KOPR agonists, while all C-13 amides were inactive. Finally, substitution of oxadiazoles for the furan ring abolished affinity for the KOPR. None of the compounds displayed any KOPR antagonism or any affinity for mu or delta opioid receptors.

In adult animals and humans, activation of kappa-opioid receptors results in a diuresis. The aim of the present study was to investigate whether kappa-opioids are also diuretic early in life and whether this is altered during postnatal maturation. Therefore, the renal effects of the kappa-opioid-receptor agonist U-50488H were measured in two separate age groups of conscious lambs at two stages of postnatal maturation (approximately 1 wk and approximately 6 wk) under physiological conditions. To evaluate whether the renal responses to U-50488H resulted from receptor-dependent effects, responses to U-50488H were also tested in the presence of the specific kappa-opioid-receptor antagonist 5'-guanidinonaltrindole (GNTI). Urinary flow rate, free water clearance, and electrolyte excretions and clearances were measured for 30 min before and for 90 min after intravenous injection of U-50488H or vehicle. An increase in urinary flow rate accompanied by an increase in free water clearance occurred in response to administration of U-50488H but not vehicle. There were no effects of U-50488H on electrolyte excretions or clearances at either 1 or 6 wk of postnatal life. Although there were no effects of GNTI on any of the measured or calculated variables, the aforementioned diuretic response to U-50488H was abolished by pretreatment with GNTI in both age groups. We conclude that kappa-opioid receptors are diuretic early in life and that this response does not appear to be altered as postnatal maturation proceeds. Therefore, these data provide evidence that activation of kappa-opioid receptors early in life may lead to alterations in fluid balance.

Rationale: Opioid receptors are divided into three types: kappa, mu, and delta receptors. Receptor-selective antagonists are useful experimental tools for evaluation of opioid receptor-mediated processes. 5′-Guanidinonaltrindole (GNTI) was recently developed as a novel kappa-selective antagonist. Objectives: To evaluate the potency, time course, and selectivity of GNTI's opioid antagonist effects in rhesus monkeys in an assay of schedule-controlled responding. Methods: Five rhesus monkeys were trained to respond under a fixed ratio 30 schedule of food reinforcement. The rate-decreasing effects of the kappa agonists U50,488 and U69,593, the mu agonist morphine, and the delta agonist SNC80 were examined alone and after pretreatment with GNTI (0.1 and 1.0 mg/kg i.m.; 1 h to 14 days). Results: U50,488, U69,593, morphine, and SNC80 dose-dependently decreased response rates in this procedure. GNTI produced a dose- and time-dependent antagonism of the rate-decreasing effects of U50,488. The kappa antagonist effects of GNTI had a slow onset and a long duration of action, and peak antagonist effects were observed after 24 h. A higher dose of 3.2 mg/kg GNTI eliminated responding in one monkey and was not studied further. The antagonist effects of GNTI were kappa selective, because 1.0 mg/kg GNTI also antagonized the effects of U69,593, but not those of morphine or SNC80. Conclusions: These results suggest that GNTI is a potent and selective kappa antagonist with a slow onset and long duration of action in rhesus monkeys. Relative to the prototype kappa antagonist nor-binaltorphimine, GNTI may have some advantages as a tool for the study of kappa receptor-mediated processes.

5′-Guanidinonaltrindole (GNTI) possesses 5-fold greater opioid antagonist potency ( K e =0.04 nM) and an order of magnitude greater selectivity (selectivity ratios >500) than the prototypical κ-opioid receptor antagonist, norbinaltorphimine, in smooth muscle preparations. Binding and functional studies conducted on cloned human opioid receptors expressed in Chinese hamster ovarian (CHO) cells afforded pA 2 values that were comparable to the smooth muscle data. In view of the high selectivity and potency of GNTI, it is a potentially valuable pharmacological tool for opioid research.

Gastrin-releasing peptide (GRP) has been implicated in the itch-scratch cycle. We investigated if this gut–brain–skin peptide plays a role in the compulsive, hindleg scratching of the neck of mice by 5′-guanidinonaltrindole (GNTI), the kappa opioid receptor antagonist, and in the antipruritic activity of nalfurafine, the kappa opioid agonist. Previously, we showed that GNTI (0.03–1 mg/kg, s.c.) elicits dose-related scratching and that nalfurafine (0.001–0.02 mg/kg, s.c.) inhibits this behavior in mice. Utilizing immunohistochemistry, GRP positive nerve fibers were detected in mouse skin and superficial layer of the dorsal horn of the spinal cord as well as GRP positive cells in the dorsal root ganglion. Pretreating mice with either a pseudopeptide GRP receptor antagonist, RC-3095 (10–30 mg/kg, s.c. at −15 min), or a peptide GRP receptor antagonist, [ d -Phe 6 ]bombesin(6-13) methyl ester (2–100 nmol, i.t. at −10 min), did not suppress GNTI-induced scratching. However, pretreating mice with either antagonist inhibited scratching precipitated by the GRP receptor agonist, GRP 18–27 (2 nmol, i.t.). Pretreating mice with a muscarinic M 1 receptor agonist, McN-A-343 (1.5–15 μg/5 μl, i.t. at −10 min) antagonized GNTI-induced scratching. Norbinaltorphimine (20 mg/kg, i.p. at −18 to −20 h), a kappa opioid antagonist, countered the antiscratch activity of nalfurafine. We conclude that (a) the GRP receptor system does not mediate GNTI-induced scratching and (b) the kappa opioid system is involved, at least in part, in the scratch suppressing activity of nalfurafine.

Background Nor-BNI, GNTI and JDTic induce κ opioid antagonism that is delayed by hours and can persist for months. Other effects are transient. It has been proposed that these drugs may be slowly absorbed or distributed, and may dissolve in cell membranes, thus slowing elimination and prolonging their effects. Recent evidence suggests, instead, that they induce prolonged desensitization of the κ opioid receptor. Methods To evaluate these hypotheses, we measured relevant physicochemical properties of nor-BNI, GNTI and JDTic, and the timecourse of brain and plasma concentrations in mice after intraperitoneal administration (using LC-MS-MS). Results In each case, plasma levels were maximal within 30 min and declined by >80% within four hours, correlating well with previously reported transient effects. A strong negative correlation was observed between plasma levels and the delayed, prolonged timecourse of κ antagonism. Brain levels of nor-BNI and JDTic peaked within 30 min, but while nor-BNI was largely eliminated within hours, JDTic declined gradually over a week. Brain uptake of GNTI was too low to measure accurately, and higher doses proved lethal. None of the drugs were highly lipophilic, showing high water solubility (> 45 mM) and low distribution into octanol (log D7.4 < 2). Brain homogenate binding was within the range of many shorter-acting drugs (>7% unbound). JDTic showed P-gp-mediated efflux; nor- BNI and GNTI did not, but their low unbound brain uptake suggests efflux by another mechanism. Conclusions The negative plasma concentration-effect relationship we observed is difficult to reconcile with simple competitive antagonism, but is consistent with desensitization. The very slow elimination of JDTic from brain is surprising given that it undergoes active efflux, has modest affinity for homogenate, and has a shorter duration of action than nor-BNI under these conditions. We propose that this persistence may result from entrapment in cellular compartments such as lysosomes.