Effects of pregabalin and duloxetine on neurotransmitters in the dorsal horn of the spinal cord in a rat model of fibromyalgia.

Dysfunction of the monoamine systems in the nervous system is associated with the clinical symptoms of fibromyalgia. Reserpine-induced myalgia (RIM) rats are a putative model of fibromyalgia in which muscle pressure thresholds and monoamine content is reduced in the brain and spinal cord. We examined the effects of pregabalin and duloxetine, drugs approved for fibromyalgia treatment, on the levels of extracellular neurotransmitters in the dorsal horn of the spinal cord in RIM rats using microdialysis. Male SD rats were used for all experiments. To generate RIM rats, reserpine was injected at 1 mg/kg subcutaneously once daily for three consecutive days. The pressure threshold of the mid-gastrocnemius muscle was measured using a Randall-Selitto apparatus. Norepinephrine, dopamine, and serotonin were detected using high-performance liquid chromatography with electrochemical detection, and glutamate and γ-aminobutyric acid (GABA) were detected using liquid chromatography-mass spectrometry. The muscle pressure threshold in RIM rats was significantly lower than that in normal rats. While the levels of monoamines and glutamate were lower in the spinal cord of RIM rats than in normal rats, levels of GABA did not markedly differ. Duloxetine increased the levels of all three monoamines in normal and RIM rats in a dose-dependent manner. In contrast, pregabalin only increased norepinephrine levels in RIM rats. These results indicate that while both pregabalin and duloxetine ameliorate muscle pressure thresholds in RIM rats, their effects on the levels of extracellular neurotransmitters in the spinal cord differ considerably.

An automated method by which effects of compounds on locomotor activity and spontaneous neuropathic pain-specific movements can be simultaneously evaluated in rats with chronic-constriction nerve injury.

Neuropathic pain patients are characterized by evoked pain (hyperalgesia and allodynia) and spontaneous pain, the latter of which is the predominant symptom. In animal models of neuropathic pain, effects of test compounds on spontaneous pain-related behaviors have been evaluated by direct visual observation. In addition, by performing another locomotor activity experiment in normal animals, it is also indispensable to examine whether test compounds cause motor impairment to avoid overestimation of their analgesic activities. In the present study, we developed spontaneous pain-specific and automated evaluation method by improving a previous method for measuring movements of the injured hind limb in unilateral chronic constriction nerve injury (CCI) rats. Rats with unilateral CCI were implanted with strong and weak magnets in each hind limb, respectively. Limb movements were automatically detected as spiked waveforms in electromagnetic field analyzing system. Movements in each limb were analyzed separately according to differences in their respective wave amplitudes, and aberrant movements of injured limb were specifically detected on basis of the asymmetry between injured and uninjured limb movements. Consequently, the incidence ratio of spontaneous pain in injured limb, which was not affected by individual locomotive activities, was able to be obtained as a new evaluation index. The incidence ratio of spontaneous pain revealed substantial difference between CCI and sham rats with only a small variation in the value. Further, according to the frequency of movement of the uninjured limb, it could be determined simultaneously whether a test compound causes sedative effect or not, resulting in eliminating the need for another experiment in normal animals.

Pharmacological characterization of a novel, potent adenosine A1 and A2A receptor dual antagonist, 5-[5-amino-3-(4-fluorophenyl)pyrazin-2-yl]-1-isopropylpyridine-2(1H)-one (ASP5854), in models of Parkinson's disease and cognition.

Central adenosine A(2A) receptor is a promising target for drugs to treat Parkinson's disease (PD), and the central blockade of adenosine A(1) receptor improves cognitive function. In the present study, we investigated the effect of a novel adenosine A(1) and A(2A) dual antagonist, 5-[5-amino-3-(4-fluorophenyl) pyrazin-2-yl]-1-isopropylpyridine-2(1H)-one (ASP5854), in animal models of PD and cognition. The binding affinities of ASP5854 for human A(1) and A(2A) receptors were 9.03 and 1.76 nM, respectively, with higher specificity and no species differences. ASP5854 also showed antagonistic action on A(1) and A(2A) agonist-induced increases of intracellular Ca(2+) concentration. ASP5854 ameliorated A(2A) agonist 2-[p-(2-carboxyethyl) phenethylamino]-5'-N-ethylcarboxamidoadenosine (CGS21680)- and haloperidol-induced catalepsy in mice, with the minimum effective doses of 0.32 and 0.1 mg/kg, respectively, and it also improved haloperidol-induced catalepsy in rats at doses higher than 0.1 mg/kg. In unilateral 6-hydroxydopamine-lesioned rats, ASP5854 significantly potentiated l-dihydroxyphenylalanine (L-DOPA)-induced rotational behavior at doses higher than 0.032 mg/kg. ASP5854 also significantly restored the striatal dopamine content reduced by 1-metyl-4-phenyl-1,2,3,6-tetrahydropyridine treatment in mice at doses higher than 0.1 mg/kg. Furthermore, in the rat passive avoidance test, ASP5854 significantly reversed the scopolamine-induced memory deficits, whereas the specific adenosine A(2A) antagonist 8-((E)-2-(3,4-dimethoxyphenyl)ethenyl)-1,3-diethyl-7-methyl-3,7-dihydro-1H-purine-2,6-dione (KW-6002; istradefylline) did not. Scopolamine- or 5H-dibenzo[a,d]cyclohepten-5,10-imine (dizocilpine maleate) (MK-801)-induced impairment of spontaneous alternation in the mouse Y-maze test was ameliorated by ASP5854, whereas KW-6002 did not exert improvement at therapeutically relevant dosages. These results demonstrate that the novel, selective, and orally active dual adenosine A(1) and A(2A) receptors antagonist ASP5854 improves motor impairments, is neuroprotective via A(2A) antagonism, and also enhances cognitive function through A(1) antagonism.

Anxiolytic activity of a novel potent serotonin 5-HT2C receptor antagonist FR260010: a comparison with diazepam and buspirone.

Hyperfunction of brain 5-hydroxytryptamine(2C) (5-HT(2C)) receptor is suggested to be involved in anxiety as evidenced by the fact that a putative 5-HT(2C) receptor agonist 1-(m-chlorophenyl)-piperazine (m-CPP) causes anxiety in humans. We have recently identified FR260010 (N-[3-(4-methyl-1H-imidazol-1-yl)phenyl]-5,6-dihydrobenzo[h]quinazolin-4-amine dimethanesulfonate) as novel 5-HT(2C) receptor antagonist from diaryl amine derivatives, and here characterized in vitro and in vivo profiles of the compound. FR260010 showed high affinity for human 5-HT(2C) receptor (K(i): 1.10 nM) and high selectivity over 5-hydroxytryptamine(2A) (5-HT(2A)) receptor (K(i): 386 nM) and many other transmitter receptors. FR260010 showed antagonist activity at human 5-HT(2C) receptor in an intracellular calcium assay and showed no detectable intrinsic activity. The compound dose-dependently inhibited the hypolocomotion (ID(50): 1.89 mg/kg, p.o.) and hypophagia (ID(50): 2.84 mg/kg, p.o.) in rats induced by m-CPP, putative indices of brain 5-HT(2C) receptor antagonist activity. We then compared the effects of FR260010 with those of two other anxiolytics belonging to different classes, diazepam and buspirone, in anxiety models in rats and mice and adverse effect tests in mice. FR260010 (0.1-3.2 mg/kg, p.o.) and diazepam (1-10 mg/kg, p.o.) decreased behavioral indices of anxiety in all models, whereas buspirone (0.32-10 mg/kg, p.o.) did not significantly affect them in any models. In adverse effect tests, FR260010 and buspirone showed modest effects, whereas diazepam showed significant effects in all tests. These results suggest that FR260010 is a novel, potent, orally active and brain penetrable antagonist of 5-HT(2C) receptor, and may have therapeutic potential for treatment of anxiety, with more desirable profiles than benzodiazepines or 5-hydroxytryptamine(1A) (5-HT(1A)) receptor agonists.