Effect Of NAN-190 Research Articles

NAN-190, a 5-HT1A antagonist, alleviates inflammatory pain by targeting Nav1.7 sodium channels

AimsIn the present study, NAN-190 [1-(2-methoxyphenyl)-4-[4-(2-phthalimido) butyl] piperazine] was identified as a Nav1.7 blocker. In the meantime, the compound could alleviate the Complete Freund's Adjuvant (CFA)-induced inflammatory pain. To understand the molecular mechanisms of NAN-190 on pain, the effect of NAN-190 on Nav1.7 sodium channels was studied. Main methodsInflammatory pain was induced by injection of CFA solution into the plantar side of the left hindpaw. Thermal hyperalgesia and mechanical allodynia were measured. Whole-cell patch clamp methods were used to record sodium channels and other pain-related targets in the cultured recombinant cells and dorsal root ganglion neurons. Key findingsNan-190 was identified as an inhibitor of Nav1.7 sodium channels and animal experiments showed that NAN-190 significantly alleviated CFA-induced inflammatory pain. Mechanism studies demonstrated that NAN-190 was a state-dependent Nav1.7 blocker with IC50 value on the inactivated state ten-fold more potent than that on the rest state. NAN-190 leftward-shifted the fast and slow inactivation curves about 9.07 mV and 38.56 mV, respectively, but had no effects on channel activation. The compound also slowed the recovery from fast and slow inactivation and showed use-dependent properties. Further, the site-directed mutagenesis experiments demonstrated that NAN-190 mainly worked on the open state of Nav1.7 channels by interacting with sites similar as local anesthetics. In DRG neurons, NAN-190 mainly blocks TTX-sensitive currents but is less sensitive to TTX-R sodium currents. SignificanceTaken together, our results indicated that NAN-190 alleviated pain behaviors by blocking sodium channels by interacting with the open state.

Effects of 8-OH-DPAT and NAN-190 on anxiety behavior, monoamine metabolism, and the level of sex hormones in female rats during the estrus cycle

In this study, we performed a comparative analysis of the effects of chronic administration of an agonist of 5-HT1A receptors, 8-OH-DPAT, (0.05 mg/kg, subcutaneously) and an antagonist of 5-HT1A receptors, NAN-190, (0.1 mg/kg, intraperitoneally) for 14 days on the anxiety behavior, monoamine metabolism in the hippocampus and amygdala, and the level of sex hormones in the blood of adult female rats during the estrus cycle. Anxiety behavior was tested in an elevated X-maze. The level of monoamines and their metabolites was evaluated using HPLC with an electrochemical detector; the content of sex hormones in the blood was measured using an enzyme immunoassay. We found that chronic administration of 8-OH-DPAT has an anxiolytic effect in rats in estrus and proestrus. After treatment with NAN-190, we found clear modulation of anxiety behavior that depended on the phase of the estrus cycle, an anxiolytic effect at a high level of endogenous estrogens and an anxiogenic effect at a low level of estrogens. The anxiolytic effect of 8-OH-DPAT in female rats was accompanied by its suppressive influence on the pituitary-ovarian and noradrenergic systems and by its activating effect on the dopaminergic system of the brain. In contrast, the behavioral effects of NAN-190 in female rats during key phases of the estrus cycle corresponded to its stimulating effect of the pituitary-ovarian system and the absence of any effect on the monoaminergic systems of the brain. These data suggest that the mechanisms of anxiety include tight interaction between the ovarian hormonal system and the serotonergic brain system.

NAN-190, a possible specific antagonist for methamphetamine

Effect of NAN-190, a selective 5-HT 1A receptor antagonist, on methamphetamine-induced locomotor activity, anorexia, analgesia, and hyperthermia was investigated in male mice. Methamphetamine (1.5 mg/kg, i.p) produced a significant increase in locomotor activity, which was significantly antagonized by NAN-190 at a dose of 4 mg/kg, i.p. NAN-190 did not alter the antinociceptive activity of mice when it was administered alone. Methamphetamine (2 mg/kg, i.p) produced a significant decrease in food intake of mice, which were deprived of food during the previous 24 h. This anorectic activity of methamphetamine was significantly antagonized by NAN-190 at a dose of 2 mg/kg, i.p. NAN-190 did not alter the food intake of mice when it was administered alone. Methamphetamine (2 mg/kg, i.p) also produced a significant increase in body temperature of mice, which was significantly antagonized by NAN-190 at a dose of 0.5 mg/kg, i.p. NAN-190 did not alter the body temperature of mice when it was administered alone. In the writhing test, methamphetamine (1 mg/kg, i.p) produced a significant antinociceptive effect in mice. This was significantly antagonized by NAN-190 at a dose of 1 mg/kg, i.p. NAN-190 did not alter the antinociceptive activity of mice when it was administered alone. The results of the present study indicate a possible role for serotonergic mechanisms, in addition to the catecholaminergic systems, in the above-studied activities of methamphetamine in mice. This role is possibly mediated through direct stimulation of the 5-HT 1A receptor subtype. All of the above-studied activities of methamphetamine were antagonized by NAN-190, which may indicate that NAN-190 is a possible antagonist for methamphetamine.

Evidence that NAN-190-induced hypotension involves vascular α 1-adrenoceptor antagonism in the rat

The effect of NAN-190 (1-(2-methoxyphenyl)-4-[4-(2-phthalimido]-butyl] piperazine), described as a mixed 5-HT 1A receptor agonist/antagonist, on cardiovascular function was studied. The i.v. injection of NAN-190 (1–300 μg/kg) dose-dependently decreased blood pressure ( p<0.001), while heart rate was not significantly modified compared to saline-treated, anaesthetized adult rats. WAY 100635 ( N-[2-[4-(2-methoxyphenyl)-1-piperazinyl] ethyl]- N-(2-pyridinyl) cyclohexanecarboxamide), a highly selective 5-HT 1A receptor antagonist, increased NAN-190-induced hypotension ( p<0.05). In the pithed rat NAN-190 displaced the phenylephrine dose–pressor response curve to the right; ED 50 values were: ≈14, 20, 40 and 270 μg/kg for saline and NAN-190 (1, 10 and 100 μg/kg, respectively); similar ED 50 values were obtained with prazosin (≈20, 69 and 358 μg/kg for 1, 10 and 100 μg/kg of prazosin, respectively). NAN-190 shifted to the right the concentration–response curves to phenylephrine in rat tail artery (α 1A-adrenoceptors), in rabbit aorta (α 1B-adrenoceptors) and in rat aorta (α 1D-adrenoceptors), with p A 2 values of 9.47, 9.02 and 9.99; while Schild slopes were −0.78, −1.13 and −0.90, respectively (not significantly different from unity). The results show that NAN-190 induced hypotension in the anaesthetized, adult rat and suggest that this effect could be explained by antagonism of vascular α 1-adrenoceptors.

Withdrawal from continuous or intermittent cocaine: Effects of NAN-190 on cocaine-induced locomotion

Rats were pretreated with 40 mg/kg/day cocaine for 14 days by either SC injections or osmotic minipumps. Rats were then withdrawn from the pretreatment regimen for 7 days. In Experiment 1, rats received 0- to 2.0-mg/kg IP injections of 1-(2-methoxyphenyl)-4[4-(2-pthalimido)butyl]piperazine (NAN-190), a putative 5-hydroxytryptamine 1A (5-HT 1A) receptor antagonist. In Experiment 2, rats received the same doses of NAN-190 in combination with a 15-mg/kg IP injection of cocaine. The results of Experiment 1 indicate that the continuous-infusion group demonstrated a dose-dependent suppression of locomotor behavior by single doses of NAN-190. NAN-190 had no consistent dose-dependent effect on the locomotor behavior of subjects in the other pretreatment groups. The results of Experiment 2 indicate that rats receiving intermittent, daily injections tended to exhibit behavior consistent with 5-HT 1A receptor supersensitivity. In contrast, rats receiving continuous cocaine tended to exhibit behavior consistent with 5-HT 1A receptor subsensitivity. Changes in 5-HT 1A receptor sensitivity may contribute to some of the anxiety and depressive symptoms exhibited by human cocaine abusers.

NAN-190: agonist and antagonist interactions with brain 5-HT 1A receptors

NAN-190 has been reported to be a 5-HT 1A antagonist in drug discrimination studies. In order to determine if the effect of NAN-190 was directly due to competitive inhibition at 5-HT 1A receptors, 5-HT 1A-mediated inhibition of adenylyl cyclase in hippocampal membranes was investigated. NAN-190 (10 −10–10 −5 M), by itself, was found to have no effect on forskolin-stimulated adenylyl cyclase. NAN-190, however, did shift the 5-carboxamidotryptamine (a 5-HT 1A agonist) log-concentration inhibition curve to the right in a concentration-dependent manner, typical of competitive antagonism. Schild analysis revealed a KB 5-HT 1A antagonist using the in vitro adenylyl cyclase system. [ 3H]NAN-190 was synthesized and its 5-HT 1A receptor binding properties were characterized and compared with the 5-HT 1A agonist radioligand, [ 3H]8-hydroxy-2-(di-n-propylamino)tetralin ([ 3H]8-OH-DPAT). The 5-HT 1A agonists, serotonin (5-HT) and 8-hydroxy-2-(di-n-propylamine(tetralin (8-OH-DPAT) competed with equal affinities regardless of the radiogland used to label the 5-HT 1A receptors. [ 3H]NAN-190 and [ 3H]8-OH-DPAT labeled the same number of sites in rat hippocampus, striatum and frontal cortex. Guanosine-5′-O-(3-thio)triphosphate (GTPγS) and 5-guanyly;-imidodiphosphate (GppNHp)m non-hydrolyzable analogs of GTP, inhibited specific [ 3H]NAN-190 binding. Adenosine-5′-O-(3-thio)triphosphate (ATPγS) and 5-adenylyl-imidodiphosphate (AppNHp) were ineffective. This guanylyl nucleotide-specific effect is generally associated with agonist radioligand binding to a GTP-binding protein coupled receptor. However, [ 3H]8-OH-DPAT was far more sensitive than [ 3H]NAN-190 to the B max reducing effects of GTP and GTPγS. We propose that the test for a reduction in B max by non-hydrolyzable guanylyl nucleotides may be more sensitive than other tests for quantifying agonist activity and may demonstrate that NAN-190 has low intrinsic activity. In summary, NAN-190 displayed antagonist-like properties in functional models of 5-HT 1A receptor activity and possibly partial agonist-like properties in radioligand binding experiments.

Mixed agonist/antagonist properties of NAN-190 at 5-HT 1A Receptors: Behavioural and in vivo brain microdialysis studies

1-(2-Methoxyphenyl)-4-[4-(2-phthalimido)butyl]piperazine, NAN-190, is a novel compound with putative 5-HT 1A antagonist properties. In the present study, the effects of NAN-190 were examined with regard to functional pre- and postsynaptic 5-HT 1A receptor-mediated events, using in vivo brain microdialysis and behavioural techniques. Our findings provide evidence that NAN-190 acts as a mixed agonist/antagonist at central 5-HT 1A receptors. Thus, NAN-190 blocked (+)8-OH-DPAT-induced behaviour in reserpinized rats, indicating antagonist properties at postsynaptic 5-HT 1A receptors. However, the compound was also able to decrease the release of 5-HT in vivo, tentatively due to an agonist action at somatodendritic 5-HT 1A autoreceptors. These data extend previous information on the pharmacological profile of NAN-190 and further emphasizes the difference between pre- and postsynaptic 5-HT 1A receptors in brain.