Characterization and comparison of rat monosodium iodoacetate and medial meniscal tear models of osteoarthritic pain.

Osteoarthritis (OA) is a degenerative form of arthritis that can result in loss of joint function and chronic pain. The pathological pain state that develops with OA disease involves plastic changes in the peripheral and central nervous systems, however, the cellular mechanisms underlying OA are not fully understood. We characterized the medial meniscal tear (MMT) surgical model and the intra-articular injection of monosodium iodoacetate (MIA) chemical model of OA in rats. Both models produced histological changes in the knee joint and associated bones consistent with OA pathology. Both models also increased p38 activation in the L3, but not L4 dorsal root ganglia (DRG), increased tyrosine hydroxylase immunostaining in the L3 DRG indicating sympathetic sprouting, and increased phosphorylated (p)CREB in thalamic neurons. In MIA-OA, but not MMT-OA rats, p38 and pERK were increased in the spinal cord, and pCREB was enhanced in the prefrontal cortex. Using in vivo electrophysiology, elevated spontaneous activity and increased responsiveness of wide dynamic range neurons to stimulation of the knee was found in both models. However, a more widespread sensitization was observed in the MIA-OA rats as neurons with paw receptive fields spontaneously fired at a greater rate in MIA-OA than MMT-OA rats. Taken together, the MIA and MMT models of OA share several common features associated with histopathology and sensitization of primary somatosensory pathways, but, observed differences between the models highlights unique consequences of the related specific injuries, and these differences should be considered when choosing an OA model and when interpreting data outcomes. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res.

Mechanistic insights into the analgesic efficacy of A-1264087, a novel neuronal Ca(2+) channel blocker that reduces nociception in rat preclinical pain models.

UNLABELLED: Voltage-gated Ca(2+) channels play an important role in nociceptive transmission. There is significant evidence supporting a role for N-, T- and P/Q-type Ca(2+) channels in chronic pain. Here, we report that A-1264087, a structurally novel state-dependent blocker, inhibits each of these human Ca(2+) channels with similar potency (IC50 = 1-2 µM). A-1264087 was also shown to inhibit the release of the pronociceptive calcitonin gene-related peptide from rat dorsal root ganglion neurons. Oral administration of A-1264087 produces robust antinociceptive efficacy in monoiodoacetate-induced osteoarthritic, complete Freund adjuvant-induced inflammatory, and chronic constrictive injury of sciatic nerve-induced, neuropathic pain models with ED50 values of 3.0, 5.7, and 7.8 mg/kg (95% confidence interval = 2.2-3.5, 3.7-10, and 5.5-12.8 mg/kg), respectively. Further analysis revealed that A-1264087 also suppressed nociceptive-induced p38 and extracellular signal-regulated kinase 1/2 phosphorylation, which are biochemical markers of engagement of pain circuitry in chronic pain states. Additionally, A-1264087 inhibited both spontaneous and evoked neuronal activity in the spinal cord dorsal horn in complete Freund adjuvant-inflamed rats, providing a neurophysiological basis for the observed antihyperalgesia. A-1264087 produced no alteration of body temperature or motor coordination and no learning impairment at therapeutic plasma concentrations. PERSPECTIVE: The present results demonstrate that the neuronal Ca(2+) channel blocker A-1264087 exhibits broad-spectrum efficacy through engagement of nociceptive signaling pathways in preclinical pain models in the absence of effects on psychomotor and cognitive function.

Biochemical and pharmacological assessment of MAP-kinase signaling along pain pathways in experimental rodent models: a potential tool for the discovery of novel antinociceptive therapeutics.

Injury to the peripheral or central nervous system can induce changes within the nervous tissues that promote a state of sensitization that may underlie conditions of pathological chronic pain. A key biochemical event in the initiation and maintenance of peripheral and central neuronal sensitization associated with chronic pain is the phosphorylation and subsequent activation of mitogen-activated protein kinases (MAPKs) and immediate early gene transcription factors, in particular cAMP-response element binding protein (CREB). In this commentary we review the preclinical data that describe anatomical and mechanistic aspects of nociceptive-induced signaling along nociceptive pathways including peripheral cutaneous axons, the dorsal root ganglia, spinal cord dorsal horn and cerebral cortex. In addition to the regional manifestation of nociceptive signaling, investigations have attempted to elucidate the cellular origin of biochemical nociceptive processing in which communication, i.e. cross-talk between neurons and glia is viewed as an essential component of pathogenic pain development. Here, we outline a research strategy by which nociceptive-induced cellular signaling in experimental pain models, specifically MAPK and CREB phosphorylation can be utilized to provide mechanistic insight into drug-target interaction along the nociceptive pathways. We describe a series of studies using nociceptive inflammatory and neuropathic pain models to investigate the effects of known pain therapeutics on nociceptive-induced biochemical signaling and present this as a complementary research strategy for assessing antinociceptive activity useful in the preclinical development of novel pain therapeutics.

α7 Nicotinic receptor agonist enhances cognition in aged 3xTg-AD mice with robust plaques and tangles.

Alzheimer disease (AD) is a progressive neurodegenerative disorder with associated memory loss, spatial disorientation, and other psychiatric problems. Cholinergic system dysfunction is an early and salient feature of AD, and enhancing cholinergic signaling with acetylcholinesterase inhibitors is currently the primary strategy for improving cognition. The beneficial effects of acetylcholinesterase inhibitors, however, are typically short-lived and accompanied by adverse effects. Recent evidence suggests that activating α7 nicotinic acetylcholine receptors (α7 nAChR) may facilitate the specific modulation of brain cholinergic signaling, leading to cognitive enhancement and possibly to amelioration of AD pathologic findings. In the present study, we determined the effect of long-term treatment with the selective α7 nAChR agonist A-582941 in aged 3xTg-AD mice with robust AD-like pathology, which is particularly significant not only because this is the only mouse model that co-develops amyloid plaques and neurofibrillary tangles but also because it enabled us to explore whether A-582941 is able to restore brain function after the severe damage associated with AD. Analysis of ß-amyloid deposits, tau phosphorylation, and inflammatory cells revealed that, overall, pathologic findings were unchanged. Rather, α7 nAChR activation induced expression of c-Fos and brain-derived neurotrophic factor and phosphorylation of cyclic adenosine monophosphate response element binding and neurotrophic tyrosine receptor kinase type 2. More important, A-582941 completely restored cognition in aged 3xTg-AD mice to the level of that in age-matched nontransgenic mice. These novel findings indicate that activating α7 nAChR is a promising treatment for cognitive impairment in AD.

Effects of the novel α7 nicotinic acetylcholine receptor agonist ABT-107 on sensory gating in DBA/2 mice: pharmacodynamic characterization.

Nicotinic acetylcholine receptor (nAChR) agonists improve sensory gating deficits in animal models and schizophrenic patients. The aim of this study was to determine whether the novel and selective α7 nAChR full agonist 5-(6-[(3R)-1-azabicyclo[2.2.2]oct-3-yloxy]pyridazin-3-yl)-1H-indole (ABT-107) improves sensory gating deficits in DBA/2 mice. Sensory gating was measured by recording hippocampal-evoked potential P20-N40 waves and determining gating test/conditioning (T/C) ratios in a paired auditory stimulus paradigm. ABT-107 at 0.1 µmol/kg (average plasma concentration of 1.1 ng/ml) significantly improved sensory gating by lowering T/C ratios during a 30-min period after administration in unanesthetized DBA/2 mice. ABT-107 at 1.0 µmol/kg was ineffective at 30 min after administration when average plasma levels were 13.5 ng/ml. However, the 1.0 µmol/kg dose was effective 180 min after administration when plasma concentration had fallen to 1.9 ng/ml. ABT-107 (0.1 µmol/kg) also improved sensory gating in anesthetized DBA/2 mice pretreated with α7 nAChR-desensitizing doses of nicotine (6.2 µmol/kg) or ABT-107 (0.1 µmol/kg) itself. Moreover, repeated b.i.d. dosing of ABT-107 (0.1 µmol/kg) was as efficacious as a single dose. The acute efficacy of ABT-107 (0.1 µmol/kg) was blocked by the nAChR antagonist methyllycaconitine, but not by the α4ß2 nAChR antagonist dihydro-ß-erythroidine. These studies demonstrate that ABT-107 improves sensory gating through the activation of nAChRs, and efficacy is sustained under conditions of repeated dosing or with prior nAChR activation with nicotine.

The novel calpain inhibitor A-705253 prevents stress-induced tau hyperphosphorylation in vitro and in vivo.

Calcium-mediated pathologic activation of the cysteine protease calpain has been linked to neurodegenerative disorders such as Alzheimer's disease (AD) through the cleavage of proteolytic substrates that negatively affect neuronal function. Hyperphosphorylation of the microtubule-associated protein tau and the subsequent aggregation of tau filaments resulting in the intracellular formation of neurofibrillary tangles are recognized as key etiological factors in AD pathology. Cyclin-dependent kinase 5 (Cdk5), a major kinase responsible for tau hyperphosphorylation in the AD brain, becomes hyperactivated through calpain-mediated cleavage-conversion of the Cdk5 regulatory protein p35 to p25. In the present study, we examined the effects of the novel small-molecule calpain inhibitor A-705253 in acute models of tau hyperphosphorylation in vitro and in vivo. In hippocampal slices in vitro, lowering medium temperature to 33 °C increased tau phosphorylation in which incubation with A-705253 blocked low temperature-induced tau phosphorylation as measured by Western blot analysis. Pentobarbital-induced hypothermia or acute systemic LPS treatment in normal mice increased tau phosphorylation in hippocampal CA3 mossy fibers, as measured by immunohistochemistry, whereas acute A-705253 pretreatment prevented the stress-induced tau hyperphosphorylation in both models. In support of a Cdk5-mediated mechanism, A-705253 administered for two weeks in the drinking water of six month-old prepathogenic 3x Tg-AD mice resulted in decreased expression of the calpain proteolytic p25 fragment. Taken together, results of these studies suggest that calpain inhibition has potential utility in reducing tau hyperphosphorylation and may represent a novel disease-modifying approach in the treatment of AD.

Effects of α7 nicotinic acetylcholine receptor agonists on antipsychotic efficacy in a preclinical mouse model of psychosis.

RATIONALE: Antipsychotics normalize responses in the DBA/2 mouse model of prepulse inhibition (PPI), a preclinical model of sensorimotor gating deficits. The α7 nicotinic acetylcholine receptor (nAChR) as a molecular target is considered an attractive approach for improvement of cognitive deficits in schizophrenia (CDS). Assessment of clinical efficacy of novel agents in CDS involves treating patients already on antipsychotic medications. OBJECTIVE: We evaluated the effects of the combination of α7 nAChR agonists ABT-107 (0.1-10.0 mg/kg i.p.), A-582941 (0.04-4.0 mg/kg i.p.), and PNU282987 (1.0-10.0 mg/kg i.p.) with risperidone (0.1-1.0 mg/kg i.p.) or haloperidol (0.3-3.0 mg/kg i.p.), representative atypical and typical antipsychotic agents in the DBA/2 mouse PPI model. The same α7 agonists were given alone or in combination with a dose of antipsychotic medication that induces a minimal level of catalepsy in rats, an assay with predictive validity for the induction of extrapyramidal symptoms. RESULTS: The α7 nAChR agonists ABT-107, A-582941, and PNU282987 had no effect in DBA/2 mouse PPI when given alone yet increased the effects of haloperidol and risperidone. The α7 nAChR agonists did not cause catalepsy in rats, nor did they enhance antipsychotic-induced catalepsy. CONCLUSIONS: When given in combination with either a typical or atypical antipsychotic, α7 nAChR agonists did not impair efficacy in the DBA/2 J mouse PPI model. The efficacy but not the motoric side effects of antipsychotics was enhanced, suggesting that adjunctive therapy of α7 nAChR agonists not only could be useful for the treatment of cognitive deficits associated with schizophrenia but also could enhance the efficacy against positive symptoms.

Monosodium iodoacetate-induced joint pain is associated with increased phosphorylation of mitogen activated protein kinases in the rat spinal cord.

BACKGROUND: Intra-articular injection of monosodium iodoacetate (MIA) in the knee joint of rats disrupts chondrocyte metabolism resulting in cartilage degeneration and subsequent nociceptive behavior that has been described as a model of osteoarthritis (OA) pain. Central sensitization through activation of mitogen activated protein kinases (MAPKs) is recognized as a pathogenic mechanism in chronic pain. In the present studies, induction of central sensitization as indicated by spinal dorsal horn MAPK activation, specifically ERK and p38 phosphorylation, was assessed in the MIA-OA model. RESULTS: Behaviorally, MIA-injected rats displayed reduced hind limb grip force 1, 2, and 3 weeks post-MIA treatment. In the same animals, activation of phospho ERK1/2 was gradually increased, reaching a significant level at post injection week 3. Conversely, phosphorylation of p38 MAPK was enhanced maximally at post injection week 1 and decreased, but remained elevated, thereafter. Double labeling from 3-wk MIA rats demonstrated spinal pERK1/2 expression in neurons, but not glia. In contrast, p-p38 was expressed by microglia and a subpopulation of neurons, but not astrocytes. Additionally, there was increased ipsilateral expression of microglia, but not astrocytes, in 3-wk MIA-OA rats. Consistent with increased MAPK immunoreactivity in the contralateral dorsal horn, mechanical allodynia to the contralateral hind-limb was observed 3-wk following MIA. Finally, intrathecal injection of the MEK1 inhibitor PD98059 blocked both reduced hind-limb grip force and pERK1/2 induction in MIA-OA rats. CONCLUSION: Results of these studies support the role of MAPK activation in the progression and maintenance of central sensitization in the MIA-OA experimental pain model.

Mice expressing the Swedish APP mutation on a 129 genetic background demonstrate consistent behavioral deficits and pathological markers of Alzheimer's disease.

Mutant Tg2576 mice which possess the human "Swedish" APP mutation have been shown to demonstrate both Abeta plaque pathology and memory deficits in behavioral tasks. These mice are routinely maintained on a mixed C57BL/6xSJL genetic background which exhibits a high frequency of retinal degeneration allele and high variability in many behavioral assays. The same APP mutation is also available maintained on a 129 genetic background, providing more genetic homogeneity, but little data are published regarding the effects of the mutation on this background. We investigated whether transgenic mice expressing the Swedish mutation on the 129 background show similar behavioral deficits and Abeta pathology as those on the mixed background. Mice on the 129 background were tested at 6-7, 11-12, or 18-19 months of age in locomotor activity, Y-maze spontaneous alternation, and contextual fear conditioning. Differences were detected between WT and Tg mice in locomotor activity at 6-7 and 18-19 months, Y-maze at 6-7 and 11-12 months, and fear conditioning at 6-7, 11-12, and 18-19 months. In contrast, Tg mice on the mixed B6/SJL background tested at 6-7 months only demonstrated significant impairment in the contextual fear conditioning assay and in the Y-maze in one of 2 cohorts tested. Despite the behavioral differences observed, similar Abeta pathology was observed between Tg mice on the two genetic backgrounds. These results indicate that mice on the 129 genetic background may generate more consistent and robust behavioral differences, providing a useful model for testing therapeutic agents for Alzheimer's disease.

Selective alpha7 nicotinic acetylcholine receptor activation regulates glycogen synthase kinase3beta and decreases tau phosphorylation in vivo.

The alpha7 nicotinic acetylcholine receptor (nAChR) plays an important role in cognitive processes and has generated recent interest as a potential drug target for treating neurodegenerative disorders such as Alzheimer's disease (AD). The property of Ca(2+) permeation associated with alpha7 nAChR agonism may lead to Ca(2+)-dependent intracellular signaling that contribute to the procognitive and neuroprotective effects that have been described with this pharmacology. In this study, we investigated whether alpha7 nAChR agonism leads to increased phosphorylation of the inhibitory regulating amino acid residue Ser-9 on GSK3beta, a major kinase responsible for tau hyperphosphorylation in AD neuropathology. Immunohistochemical analysis revealed that the selective alpha7 agonist A-582941 increased S(9)-GSK3beta phosphorylation in mouse cingulate cortex and hippocampus that was not observed in alpha7 nAChR knock-out mice. A-582941 steady state exposure through continuous (2 wk) infusion also increased S(9)-GSK3beta phosphorylation in the hippocampus of Tg2576 (APP), as well as wild-type mice. Moreover, A-582941 continuous infusion decreased phosphorylation of tau in hippocampal CA3 Mossy fibers and spinal motoneurons in a hypothermia-induced tau hyperphosphorylation mouse model and AD double transgenic APP/tau mouse line, respectively. These studies demonstrate that inactivation of GSK3beta may be associated with alpha7 nAChR-induced signaling leading to attenuated tau hyperphosphorylation, raising the intriguing possibility that alpha7 nAChR agonism may have disease modifying benefit in the treatment of tauopathies, in particular AD.

Localization of histamine H4 receptors in the central nervous system of human and rat.

Existing data on the expression of H(4) histamine receptor in the CNS are conflicting and inconclusive. In this report, we present the results of experiments that were conducted in order to elucidate H(4) receptor expression and localization in the brain, spinal cord, and dorsal root ganglia (DRG). Here we show that transcripts of H(4) receptor are present in all analyzed regions of the human CNS, including spinal cord, hippocampus, cortex, thalamus and amygdala, with the highest levels of H(4) mRNA detected in the spinal cord. In rat, H(4) mRNA was detected in cortex, cerebellum, brainstem, amygdala, thalamus and striatum. Very low levels of H(4) mRNA were detected in hypothalamus, and no H(4) signal was detected in the rat hippocampus. Fairly low levels of H(4) mRNA were detected in examined peripheral tissues including spleen and liver. Interestingly, strong expression of H(4) mRNA was detected in the rat DRG and spinal cord. Immunohistochemical analysis revealed expression of H(4) receptors on neurons in the rat lumbar DRG and in the lumbar spinal cord. Our observations provide evidence of the H(4) presence in both human and rodent CNS and offer some insight into possible role of H(4) in itch and pain.

The novel calpain inhibitor A-705253 potently inhibits oligomeric beta-amyloid-induced dynamin 1 and tau cleavage in hippocampal neurons.

We have previously shown that beta-amyloid (Abeta) oligomers induced dynamin 1 and tau cleavage in cultured hippocampal neurons. As a result of this cleavage, dynamin 1 levels decreased and a toxic tau fragment was generated. Abeta-induced cleavage of these proteins was calpain-mediated and impacted both synaptic vesicle recycling and the integrity of neuronal processes [Kelly, B.L., Vassar, R., Ferreira, A., 2005. Beta-amyloid-induced dynamin 1 depletion in hippocampal neurons. A potential mechanism for early cognitive decline in Alzheimer disease. J. Biol. Chem. 280, 31746-31753; Park, S.Y., Ferreira, A., 2005. The generation of a 17kDa neurotoxic fragment: an alternative mechanism by which tau mediates beta-amyloid-induced neurodegeneration. J. Neurosci. 25, 5365-5375; Kelly, B.L., Ferreira, A., 2006. Beta-amyloid-induced dynamin 1 degradation is mediated by N-methyl-d-aspartate receptors in hippocampal neurons. J. Biol. Chem. 281, 28079-28089, Kelly, B.L., Ferreira, A., 2007. Beta-amyloid disrupted synaptic vesicle endocytosis in cultured hippocampal neurons. Neuroscience 147, 60-70]. Building on previous reports, these results identified calpain as a potential target for therapeutic intervention in Alzheimer's disease. In the present study, we tested the ability of A-705253, a novel water-soluble calpain inhibitor with oral availability and enhanced metabolic stability, to prevent Abeta-induced dynamin 1 and tau cleavage in cultured hippocampal neurons. Quantitative Western blot analysis indicated that the incubation of these cells with A-705253 prior to the addition of oligomeric Abeta reduced both dynamin 1 and tau cleavage in a dose-dependent manner. In addition, our results showed that this calpain inhibitor significantly ameliorated the cleavage of these proteins when added simultaneously with oligomeric Abeta. Furthermore, our data indicated that the use of this calpain inhibitor could have some beneficial effects even when added after the cleavage of these proteins have been triggered by Abeta. Collectively, these results suggest that, indeed, specific calpain inhibitors could play an important role in the treatment of Alzheimer's disease.

Broad-spectrum efficacy across cognitive domains by alpha7 nicotinic acetylcholine receptor agonism correlates with activation of ERK1/2 and CREB phosphorylation pathways.

The alpha7 nicotinic acetylcholine receptor (nAChR) plays an important role in cognitive processes and may represent a drug target for treating cognitive deficits in neurodegenerative and psychiatric disorders. In the present study, we used a novel alpha7 nAChR-selective agonist, 2-methyl-5-(6-phenyl-pyridazin-3-yl)-octahydro-pyrrolo[3,4-c]pyrrole (A-582941) to interrogate cognitive efficacy, as well as examine potential cellular mechanisms of cognition. Exhibiting high affinity to native rat (Ki = 10.8 nM) and human (Ki = 16.7 nM) alpha7 nAChRs, A-582941 enhanced cognitive performance in behavioral assays including the monkey delayed matching-to-sample, rat social recognition, and mouse inhibitory avoidance models that capture domains of working memory, short-term recognition memory, and long-term memory consolidation, respectively. In addition, A-582941 normalized sensory gating deficits induced by the alpha7 nAChR antagonist methyllycaconitine in rats, and in DBA/2 mice that exhibit a natural sensory gating deficit. Examination of signaling pathways known to be involved in cognitive function revealed that alpha7 nAChR agonism increased extracellular-signal regulated kinase 1/2 (ERK1/2) phosphorylation in PC12 cells. Furthermore, increases in ERK1/2 and cAMP response element-binding protein (CREB) phosphorylation were observed in mouse cingulate cortex and/or hippocampus after acute A-582941 administration producing plasma concentrations in the range of alpha7 binding affinities and behavioral efficacious doses. The MEK inhibitor SL327 completely blocked alpha7 agonist-evoked ERK1/2 phosphorylation. Our results demonstrate that alpha7 nAChR agonism can lead to broad-spectrum efficacy in animal models at doses that enhance ERK1/2 and CREB phosphorylation/activation and may represent a mechanism that offers potential to improve cognitive deficits associated with neurodegenerative and psychiatric diseases, such as Alzheimer's disease and schizophrenia.

Antisense knockdown of the rat alpha7 nicotinic acetylcholine receptor produces spatial memory impairment.

Selective and brain penetrating pharmacological antagonists for use in clarifying a role of alpha7 nicotinic acetylcholine receptors (nAChR) in behavioral paradigms are presently unavailable. Studies in alpha7 knock-out mice (KO) have not revealed convincing changes in behavioral phenotype, in particular measures of cognition that include contextual fear conditioning and spatial memory, which may be due to compensatory developmental changes. Therefore, an antisense oligonucleotide (aON) targeted toward the 3'- and 5'-UTR coding regions of the rat alpha7 nicotinic acetylcholine receptor was used. Following central injection of aON into the lateral ventricle of Long Evans rats for 6 days, treated rats exhibited a significant 42% and 25% decrease in alpha7 nAChR densities in hippocampus and cortex, respectively, as measured by [(3)H]-methyllycaconitine (MLA) binding. There was no change in alpha4beta2 densities measured by [(3)H]-cytisine binding. Acquisition of Morris Water Maze (MWM) performance, a measure of spatial memory, was impaired in aON-treated rats. In addition, a reduction in target platform crossings during a subsequent probe-trial was observed. These data demonstrate the ability of this aON to reduce hippocampal and cortical alpha7 nicotinic receptor densities associated with impaired MWM performance and support the specific involvement of the alpha7 nAChR in spatial learning and memory, a phenotype not affected in alpha7 KO mice.

A-85380: a pharmacological probe for the preclinical and clinical investigation of the alphabeta neuronal nicotinic acetylcholine receptor.

A-85380 [3-(2(s)-azetidinylmethoxy) pyridine] is a neuronal nicotinic acetylcholine receptor (nAChR) agonist that has been a useful tool in the investigation of the function of nAChRs in both preclinical and clinical studies. Amongst nAChR subtypes, A-85380 shows selectivity for the alpha(4)beta(2) vs. the alpha(7) or alpha(1)beta(1)deltagamma nAChRs. In functional in vitro cation flux assays, A-85380 is a potent and full agonist. A-85380 has a broad-spectrum analgesic profile with efficacy in acute, persistent, and neuropathic pain models. As demonstrated using selective nAChR antagonists or alpha(4) antisense, the alpha(4)beta(2) nAChR mediates the analgesic effects of A-85380. Interestingly, the site of action depends upon the type of pain as antinociception is mediated by descending inhibition into the spinal cord whereas anti-allodynia in neuropathic pain is mediated at both central and peripheral sites. Radiolabelled forms of A-85380 have been developed and shown to be safe for use in vivo in humans. In clinical studies using positron and photon emission tomography, marked decreases in alpha(4)beta(2) nAChRs have been seen in patients with Parkinson's and Alzheimer's disease. Although not developed as a therapeutic agent, A-85380 has proven to be an important component in the development of novel nAChR ligands for the treatment of pain and other disorders.

Alpha4beta2 nicotinic receptor stimulation contributes to the effects of nicotine in the DBA/2 mouse model of sensory gating.

RATIONALE: Nicotine improves the deficiencies of sensory gating function in schizophrenic patients and in dilute brown non-Agouti (DBA/2) mice. This effect of nicotine has been attributed to activation of the alpha7 nicotinic acetylcholine receptor (nAChR) subtype. OBJECTIVE: The aim of this study was to determine whether the activation of another nAChR subtype, the central nervous system (CNS) prominent alpha4beta2 receptor, also contributes to the effects of nicotine on sensory gating in DBA/2 mice. METHODS: Unanesthetized DBA/2 mice were treated either with nicotine, the alpha4beta2 antagonist dihydro-beta-erythroidine, the noncompetitive nAChR antagonist mecamylamine, or a combination of an antagonist and nicotine. Thereafter, gating was assessed by recording hippocampal evoked potentials (EP), which were elicited by pairs of auditory clicks. The EP response to the second click, or test amplitude (TAMP), was divided by the EP response to the first click, or condition amplitude (CAMP), to derive gating T:C ratios. RESULTS: Nicotine significantly (p<0.05) lowered T:C ratios by 42%, while significantly increasing CAMP by 55%. After a pretreatment with dihydro-beta-erythroidine, nicotine still significantly lowered T:C ratios by 28%; however, the nicotine-induced increase of CAMP was blocked. Mecamylamine blocked the effect of nicotine on both T:C ratios and CAMP. CONCLUSIONS: Activation of alpha4beta2 receptors by nicotine increases CAMP. However, under conditions where alpha4beta2 receptors are blocked, nicotine still lowers T:C ratios and may improve sensory gating, possibly through the activation of other nAChR subtypes such as alpha7. These effects of nicotine on auditory EPs may be indicative of a profile that would improve information processing in schizophrenia and other CNS diseases.

Dissociation between post-surgical pain behaviors and spinal Fos-like immunoreactivity in the rat.

Previous studies have demonstrated that Fos-like immunoreactivity is increased in spinal dorsal horn neurons in several pain models, and have suggested that Fos-like immunoreactivity could be used as a marker of neurons activated by painful stimulation. In the present study, we evaluated nociceptive behaviors and spinal Fos-like immunoreactivity in a rat skin incision model of post-operative pain. In this model, evoked and non-evoked pain behaviors were observed at least for 2 days after paw surgery, an increased number of Fos-like immunoreactive neurons was observed in the spinal dorsal horn at lumbar levels 4-5 two-hour post-surgery. The number of Fos-like immunoreactive neurons was significantly greater in animals with skin-muscle incision compared to animals with skin-alone incision. Interestingly, spinal Fos-like immunoreactivity was quickly normalized in rats with paw surgery at later time points (8 and 24 h post-surgery), whereas nociceptive behaviors were still observed. Furthermore, at 24 h post-surgery, spinal Fos-like immunoreactivity induced by thermal stimulation (42, 44, 46, 48, 52 degrees C for 15 s) was not significantly different between sham animals and animals with surgery. In both groups, an increase in spinal Fos-like immunoreactive neurons was observed with increasing temperatures, with similar laminar distribution. Finally, systemic morphine reduced post-operative pain and Fos-like immunoreactivity in a naloxone reversible manner, with greater potency and efficacy on behavioral endpoints than on Fos-like immunoreactivity. These results demonstrate a different profile of nociceptive behaviors and spinal Fos-like immunoreactivity in the rat skin incision model, suggesting a limited potential of spinal Fos-like immunoreactivity to study post-surgical pain and its pharmacology.

Dopamine D4 receptor signaling in the rat paraventricular hypothalamic nucleus: Evidence of natural coupling involving immediate early gene induction and mitogen activated protein kinase phosphorylation.

The dopamine D4 receptor has been investigated for its potential role in several CNS disorders, notably schizophrenia and more recently, erectile dysfunction. Whereas studies have investigated dopamine D4 receptor-mediated signaling in vitro, there have been few, if any, attempts to identify dopamine D4 receptor signal transduction pathways in vivo. In the present studies, the selective dopamine D4 agonist PD168077 induces c-Fos expression and extracellular signal regulated kinase (ERK) phosphorylation in the hypothalamic paraventricular nucleus (PVN), a site known to regulate proerectile activity. The selective dopamine D4 receptor antagonist A-381393 blocked both c-Fos expression and ERK1/2 phosphorylation produced by PD168077. In addition, PD168077-induced ERK1/2 phosphorylation was prevented by SL327, an inhibitor of ERK1/2 phosphorylation. Interestingly, treatment with A-381393 alone significantly reduced the amount of Fos immunoreactivity as compared to basal expression observed in vehicle-treated controls. Dopamine D4 receptor and c-Fos coexpression in the PVN was observed using double immunohistochemical labeling, suggesting that PD168077-induced signaling may result from direct dopamine D4 receptor activation. Our results demonstrate functional dopamine D4 receptor expression and natural coupling in the PVN linked to signal transduction pathways that include immediate early gene and MAP kinase activation. Further, the ability of the selective dopamine D4 antagonist A-381393 alone to reduce c-Fos expression below control levels may imply the presence of a tonic dopamine D4 receptor activation under basal conditions in vivo. These findings provide additional evidence that the PVN may be a site of dopamine D4 receptor-mediated proerectile activity.

Globular amyloid beta-peptide oligomer - a homogenous and stable neuropathological protein in Alzheimer's disease.

Amyloid beta-peptide (Abeta)(1-42) oligomers have recently been discussed as intermediate toxic species in Alzheimer's disease (AD) pathology. Here we describe a new and highly stable Abeta(1-42) oligomer species which can easily be prepared in vitro and is present in the brains of patients with AD and Abeta(1-42)-overproducing transgenic mice. Physicochemical characterization reveals a pure, highly water-soluble globular 60-kDa oligomer which we named 'Abeta(1-42) globulomer'. Our data indicate that Abeta(1-42) globulomer is a persistent structural entity formed independently of the fibrillar aggregation pathway. It is a potent antigen in mice and rabbits eliciting generation of Abeta(1-42) globulomer-specific antibodies that do not cross-react with amyloid precursor protein, Abeta(1-40) and Abeta(1-42) monomers and Abeta fibrils. Abeta(1-42) globulomer binds specifically to dendritic processes of neurons but not glia in hippocampal cell cultures and completely blocks long-term potentiation in rat hippocampal slices. Our data suggest that Abeta(1-42) globulomer represents a basic pathogenic structural principle also present to a minor extent in previously described oligomer preparations and that its formation is an early pathological event in AD. Selective neutralization of the Abeta globulomer structure epitope is expected to have a high potential for treatment of AD.

Preclinical profiling and safety studies of ABT-769: a compound with potential for broad-spectrum antiepileptic activity.

PURPOSE: The objective of this study was to characterize the antiseizure and safety profiles of ABT-769 [(R)-N-(2 amino-2-oxoethyl)spiro[2,5]octane-1-carboxamide]. METHODS: ABT-769 was tested for protection against maximal electroshock and pentylenetetrazol-induced seizures in the mouse and for suppression of electrically kindled amygdala seizures and spontaneous absence-like seizures in the rat. The central nervous system safety profile was evaluated by using tests of motor coordination and inhibitory avoidance. The potential for liver toxicity was assessed in vitro by using a mitochondrial fatty acid beta-oxidation assay. Teratogenic potential was assessed in the mouse. RESULTS: ABT-769 blocked maximal electroshock, subcutaneous pentylenetetrazol and intravenous pentylenetetrazol-induced seizures with median effective dose (ED50) values of 0.25, 0.38, and 0.11 mmol/kg, p.o., respectively. No tolerance was evident in the intravenous pentylenetetrazol test after twice-daily dosing of ABT-769 (0.3 mmol/kg, p.o.) for 4 days. ABT-769 blocked absence-like spike-wave discharge (ED50, 0.15 mmol/kg, p.o.) and shortened the cortical and amygdala afterdischarge duration of kindled seizures (1 and 3 mmol/kg, p.o.). The protective indices (ED50 rotorod impairment/ED50 seizure protection) were 4.8, 3.2, and 10.9 in the maximal electroshock, subcutaneous pentylenetetrazol and intravenous pentylenetetrazol seizure tests, respectively. ABT-769 did not affect inhibitory avoidance performance (0.1-1 mmol/kg, p.o.). ABT-769 did not affect mitochondrial fatty acid beta-oxidation or induce neural tube defects. CONCLUSIONS: ABT-769 is an efficacious antiseizure agent in animal models of convulsive and nonconvulsive epilepsy and has a favorable safety profile. ABT-769 has a broad-spectrum profile like that of valproic acid. Its profile is clearly different from those of carbamazepine, phenytoin, lamotrigine, topiramate, vigabatrin, and tiagabine.