Sheree F. Logue

Assessment of learning by the morris water task and fear conditioning in inbred mouse strains and F1 hybrids : implications of genetic background for single gene mutations and quantitative trait loci analyses

Genetic methods including the creation of transgenic or null mutant models and mapping studies using quantitative trait loci strategies can be used to identify candidate genes in mice that regulate learning processes. Interpretations as to the impact of single gene mutations for polygenic behaviours like learning will depend in part on the genetic background of the animals used for these manipulations. To address the issue of genetic variability, 12 inbred strains and seven different F1 hybrids were tested on multiple behavioural tasks, including two complex learning paradigms: the Morris water task and fear conditioning. Strain differences were found for all variables measured. In the hidden platform version of the Morris task, the albino animals performed poorly while overall the F1 hybrids showed the best selectivity for the trained quadrant as measured in a probe trial. In contrast, almost all genotypes performed well on the contextual fear conditioning task and learned to associate the test context with the pairing of a foot shock and auditory stimulus as demonstrated 24 h after training by increased freezing in the test environment compared to an altered context. Significant genetic correlations were obtained for behavioural measures suggesting that the same genes regulate various aspects of performance on behavioural tasks. Scores from these multiple inbred strains and F1 hybrids provide a baseline level of learning ability for fear conditioning and the Morris water task. The results of the present study confirm the importance of genetic background in the performance of various learning tasks. This variability should be considered when developing new transgenic or null mutant animal models.

Assessment of locomotor activity, acoustic and tactile startle, and prepulse inhibition of startle in inbred mouse strains and F1 hybrids: Implications of genetic background for single gene and quantitative trait loci analyses

As the use of transgenic and null mutation techniques in the development of animal models of disorders increases, the importance of selecting the appropriate genetic background also increases. The genetic background of the mouse strains used as models for various disorders is critical because of the potential for epistatic effects on the expression of transgenes and null mutations. Twelve strains of inbred mice and seven F1 hybrids were tested in multiple behavioural tasks including open-field locomotor activity, Y-maze activity, auditory and tactile startle and prepulse inhibition of startle response. Differences across genotypes were found for all variables measured. The range of variability among genotypes was dependent on the specific measure so careful consideration must be made in selecting a strain for testing a particular behaviour. Because of the polygenic nature of each of the behavioural phenotypes, the impact of a single gene manipulation may vary depending on the genetic background on which it is expressed. Moreover, quantitative trait loci methods could be applied to these behaviours.

Phosphodiesterase 10A inhibitor activity in preclinical models of the positive, cognitive, and negative symptoms of schizophrenia.

Following several recent reports that suggest that dual cAMP and cGMP phosphodiesterase 10A (PDE10A) inhibitors may present a novel mechanism to treat positive symptoms of schizophrenia, we sought to extend the preclinical characterization of two such compounds, papaverine [1-(3,4-dimethoxybenzyl)-6,7-dimethoxyisoquinoline] and MP-10 [2-{[4-(1-methyl-4-pyridin-4-yl-1H-pyrazol-3-yl)phenoxy]methyl}quinoline], in a variety of in vivo and in vitro assays. Both of these compounds were active in a range of antipsychotic models, antagonizing apomorphine-induced climbing in mice, inhibiting conditioned avoidance responding in both rats and mice, and blocking N-methyl-d-aspartate antagonist-induced deficits in prepulse inhibition of acoustic startle response in rats, while improving baseline sensory gating in mice, all of which strengthen previously reported observations. These compounds also demonstrated activity in several assays intended to probe negative symptoms and cognitive deficits, two disease domains that are underserved by current treatments, with both compounds showing an ability to increase sociality in BALB/cJ mice in the social approach/social avoidance assay, enhance social odor recognition in mice and, in the case of papaverine, improve novel object recognition in rats. Biochemical characterization of these compounds has shown that PDE10A inhibitors modulate both the dopamine D1-direct and D2-indirect striatal pathways and regulate the phosphorylation status of a panel of glutamate receptor subunits in the striatum. It is striking that PDE10A inhibition increased the phosphorylation of the (±)-α-amino-3-hydroxy-5-methylisoxazole-4-proprionic acid receptor GluR1 subunit at residue serine 845 at the cell surface. Together, our results suggest that PDE10A inhibitors alleviate both dopaminergic and glutamatergic dysfunction thought to underlie schizophrenia, which may contribute to the broad-spectrum efficacy.

ADX47273 [S-(4-Fluoro-phenyl)-{3-[3-(4-fluoro-phenyl)-[1,2,4]-oxadiazol-5-yl]-piperidin-1-yl}-methanone]: A Novel Metabotropic Glutamate Receptor 5-Selective Positive Allosteric Modulator with Preclinical Antipsychotic-Like and Procognitive Activities

Positive allosteric modulators (PAMs) of metabotropic glutamate receptor subtype 5 (mGlu5) enhance N-methyl-d-aspartate receptor function and may represent a novel approach for the treatment of schizophrenia. ADX47273 [S-(4-fluoro-phenyl)-{3-[3-(4-fluoro-phenyl)-[1,2,4]oxadiazol-5-yl]-piperidin-1-yl}-methanone], a recently identified potent and selective mGlu5 PAM, increased (9-fold) the response to threshold concentration of glutamate (50 nM) in fluorometric Ca2+ assays (EC50 = 170 nM) in human embryonic kidney 293 cells expressing rat mGlu5. In the same system, ADX47273 dose-dependently shifted mGlu5 receptor glutamate response curve to the left (9-fold at 1 μM) and competed for binding of [3H]2-methyl-6-(phenylethynyl)pyridine (Ki = 4.3 μM), but not [3H]quisqualate. In vivo, ADX47273 increased extracellular signal-regulated kinase and cAMP-responsive element-binding protein phosphorylation in hippocampus and prefrontal cortex, both of which are critical for glutamate-mediated signal transduction mechanisms. In models sensitive to antipsychotic drug treatment, ADX47273 reduced rat-conditioned avoidance responding [minimal effective dose (MED) = 30 mg/kg i.p.] and decreased mouse apomorphine-induced climbing (MED = 100 mg/kg i.p.), with little effect on stereotypy or catalepsy. Furthermore, ADX47273 blocked phencyclidine, apomorphine, and amphetamine-induced locomotor activities (MED = 100 mg/kg i.p.) in mice and decreased extracellular levels of dopamine in the nucleus accumbens, but not in the striatum, in rats. In cognition models, ADX47273 increased novel object recognition (MED = 1 mg/kg i.p.) and reduced impulsivity in the five-choice serial reaction time test (MED = 10 mg/kg i.p.) in rats. Taken together, these effects are consistent with the hypothesis that allosteric potentiation of mGlu5 may provide a novel approach for development of antipsychotic and procognitive agents.

WAY-163909 [(7bR,10aR)-1,2,3,4,8,9,10,10a-octahydro-7bH-cyclopenta-[b][1,4]diazepino[6,7,1hi]indole]: A novel 5-hydroxytryptamine 2C receptor-selective agonist with preclinical antipsychotic-like activity.

Serotonin-2C (5-HT2C) receptor antagonists and agonists have been shown to affect dopamine (DA) neurotransmission, with agonists selectively decreasing mesolimbic DA. As antipsychotic efficacy is proposed to be associated with decreased mesolimbic DA neurotransmission by virtue of DA D2 receptor antagonism, the 5-HT2C-selective receptor agonist, WAY-163909 [(7bR,10aR)-1,2, 3,4,8,9,10,10a-octahydro-7bH-cyclopenta-[b][1,4]diazepino[6,7, 1hi]indole], was evaluated in animal models of schizophrenia and in vivo microdialysis and electrophysiology to determine the effects on mesolimbic and nigrostriatal DA neurotransmission. Similar to clozapine, WAY-163909 (1.7–30 mg/kg i.p.) decreased apomorphine-induced climbing with little effect on stereotypy and no significant induction of catalepsy. WAY-163909 (0.3–3 mg/kg s.c.) more potently reduced phencyclidine-induced locomotor activity compared with d-amphetamine with no effect on spontaneous activity. WAY-163909 (1.7–17 mg/kg i.p.) reversed MK-801 (5H-dibenzo[a,d]cyclohepten-5,10-imine (dizocilpine maleate)- and DOI [1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane]-disrupted prepulse inhibition of startle (PPI) and improved PPI in DBA/2N mice. In conditioned avoidance responding, WAY-163909 (0.3–3 mg/kg i.p.; 1–17 mg/kg p.o.) reduced avoidance responding, an effect blocked by the 5-HT2B/2C receptor antagonist SB 206553 [5-methyl-1-(3-pyridylcarbamoyl)-1,2,3,5-tetrahydropyrrolo[2,3-f]indole]. WAY-163909 (10 mg/kg s.c.) selectively decreased extracellular levels of DA in the nucleus accumbens without affecting the striatum. Likewise, in vivo electrophysiological recordings showed a decrease in the number of spontaneously firing DA neurons in the ventral tegmental area but not in the substantia nigra with both acute and chronic (21-day) administration of WAY-163909 (1–10 mg/kg i.p.). Thus, the profile of the 5-HT2C selective receptor agonist WAY-163909 is similar to that of an atypical antipsychotic and additionally may have rapid onset properties.

ADX47273: A Novel Metabotropic Glutamate Receptor 5 Selective Positive Allosteric Modulator with Preclinical Antipsychotic-Like and Pro-cognitive Activities

Positive allosteric modulators (PAMs) of metabotropic glutamate receptor subtype 5 (mGlu5) enhance N-methyl-d-aspartate receptor function and may represent a novel approach for the treatment of schizophrenia. ADX47273 [S-(4-fluoro-phenyl)-{3-[3-(4-fluoro-phenyl)-[1,2,4]oxadiazol-5-yl]-piperidin-1-yl}-methanone], a recently identified potent and selective mGlu5 PAM, increased (9-fold) the response to threshold concentration of glutamate (50 nM) in fluorometric Ca2+ assays (EC50 = 170 nM) in human embryonic kidney 293 cells expressing rat mGlu5. In the same system, ADX47273 dose-dependently shifted mGlu5 receptor glutamate response curve to the left (9-fold at 1 μM) and competed for binding of [3H]2-methyl-6-(phenylethynyl)pyridine (Ki = 4.3 μM), but not [3H]quisqualate. In vivo, ADX47273 increased extracellular signal-regulated kinase and cAMP-responsive element-binding protein phosphorylation in hippocampus and prefrontal cortex, both of which are critical for glutamate-mediated signal transduction mechanisms. In models sensitive to antipsychotic drug treatment, ADX47273 reduced rat-conditioned avoidance responding [minimal effective dose (MED) = 30 mg/kg i.p.] and decreased mouse apomorphine-induced climbing (MED = 100 mg/kg i.p.), with little effect on stereotypy or catalepsy. Furthermore, ADX47273 blocked phencyclidine, apomorphine, and amphetamine-induced locomotor activities (MED = 100 mg/kg i.p.) in mice and decreased extracellular levels of dopamine in the nucleus accumbens, but not in the striatum, in rats. In cognition models, ADX47273 increased novel object recognition (MED = 1 mg/kg i.p.) and reduced impulsivity in the five-choice serial reaction time test (MED = 10 mg/kg i.p.) in rats. Taken together, these effects are consistent with the hypothesis that allosteric potentiation of mGlu5 may provide a novel approach for development of antipsychotic and procognitive agents.

Effects of atomoxetine and methylphenidate on attention and impulsivity in the 5-choice serial reaction time test.

Deficits in attention and response inhibition are apparent across several neurodegenerative and neuropsychiatric disorders for which current pharmacotherapy is inadequate. The 5-choice serial reaction time test (5-CSRTT), which originated from the continuous performance test (CPT) in humans, may serve as a useful translational assay for efficacy in these key behavioral domains. The selective norepinepherine reuptake inhibitor, atomoxetine, represents the first non-stimulant based drug approved for Attention Deficit Hyperactivity Disorder (ADHD) and has replaced methylphenidate (Ritalin) as the first line in pharmacotherapy for the treatment of ADHD. Methylphenidate and atomoxetine have different cortical and sub-cortical neurochemical signatures that could predict differences in cognitive and non-cognitive functions. The present experiments investigated the effects of acute methylphenidate and atomoxetine in male long Evans rats in the 5-choice serial reaction time (5CSRT) test that is hypothesized to serve as a model of vigilance and impulsivity behaviors associated with ADHD. Long Evans rats were trained to perform at 75% correct responses with fewer than 20% missed trials in the 5CSRT test (500 ms stimulus duration, 5 s inter-trial interval (ITI)). By varying the ITI (10, 7, 5, and 4 s) on drug test days, impulsivity (as defined by premature responses) was dramatically increased with a concomitant decrease in attention (percent correct). Subsequently, animals were treated with methylphenidate (2.5 and 5 mg/kg, i.p.) or atomoxetine (0.1, 0.5 and 1 mg/kg, i.p.) using this design. In Experiment 1, treatment with methylphenidate modestly improved overall attention but the highest dose of methylphenidate (5.0 mg/kg) significantly increased impulsivity. In contrast, treatment with atomoxetine induced a marked decrease in impulsivity whilst modestly improving overall attention. Interestingly, no effect was observed on measures of performance (e.g. motivation/sedation) with atomoxetine, whilst moderate hyperactivity (faster overall response latencies; magazine, correct, incorrect) was observed in the methylphenidate group. Those data suggest that the 5CSRT test can be used to differentiate stimulant and non-stimulant pharmacotherapies on measures of impulsivity.

Contextual and cued fear conditioning in C57BL/6J and DBA/2J mice: context discrimination and the effects of retention interval.

Context discrimination and time course studies of contextual fear conditioning revealed strain differences between C57BL/6J (B6) and DBA/2J (D2) mice. Both strains discriminated contexts, but D2 mice exhibited less freezing in a shock-paired context. The strains did not differ immediately, or at 1 and 3 hr after contextual fear conditioning training. D2 mice showed less freezing at 15 min, 30 min, and 24 hr after training. B6 mice exhibited exaggerated generalized freezing and poor discrimination between the context and altered context 7-30 days after training. The acoustic startle response in B6 mice was also enhanced at 14 days after training. D2 mice did not show this pattern of generalized freezing. B6, but not D2, mice retained contextual memories for at least 60 days.

Phosphodiesterase 11A in brain is enriched in ventral hippocampus and deletion causes psychiatric disease-related phenotypes

Phosphodiesterase 11A (PDE11A) is the most recently identified family of phosphodiesterases (PDEs), the only known enzymes to break down cyclic nucleotides. The tissue expression profile of this dual specificity PDE is controversial, and little is understood of its biological function, particularly in the brain. We seek here to determine if PDE11A is expressed in the brain and to understand its function, using PDE11A−/− knockout (KO) mice. We show that PDE11A mRNA and protein are largely restricted to hippocampus CA1, subiculum, and the amygdalohippocampal area, with a two- to threefold enrichment in the ventral vs. dorsal hippocampus, equal distribution between cytosolic and membrane fractions, and increasing levels of protein expression from postnatal day 7 through adulthood. Interestingly, PDE11A KO mice show subtle psychiatric-disease–related deficits, including hyperactivity in an open field, increased sensitivity to the glutamate N-methyl-D-aspartate receptor antagonist MK-801, as well as deficits in social behaviors (social odor recognition memory and social avoidance). In addition, PDE11A KO mice show enlarged lateral ventricles and increased activity in CA1 (as per increased Arc mRNA), phenotypes associated with psychiatric disease. The increased sensitivity to MK-801 exhibited by PDE11A KO mice may be explained by the biochemical dysregulation observed around the glutamate α-amino-3-hydroxy-5-methyl-4-isozazolepropionic (AMPA) receptor, including decreased levels of phosphorylated-GluR1 at Ser845 and the prototypical transmembrane AMPA-receptor–associated proteins stargazin (γ2) and γ8. Together, our data provide convincing evidence that PDE11A expression is restricted in the brain but plays a significant role in regulating brain function.

WAY-163909, a 5-HT2C agonist, enhances the preclinical potency of current antipsychotics

Introduction5-HT2C agonists, by decreasing mesolimbic dopamine without affecting nigrostriatal dopamine, are predicted to have antipsychotic efficacy with low extrapyramidal side effects (EPS). Combining 5-HT2C agonists with low doses of existing antipsychotics could increase treatment efficacy while reducing treatment liabilities such as EPS (typical antipsychotics), and the propensity for weight gain (atypical antipsychotics).ObjectivesThe objectives of these studies were to combine WAY-163909, a selective 5-HT2C agonist, with either the typical antipsychotic haloperidol, or the atypical antipsychotic clozapine, at doses that were ineffective on their own, with the expectation that a shift in potency in several rodent behavior models predictive of antipsychotic activity would occur.Results and discussionIn mice, co-administration of either haloperidol, or clozapine, produced a significant leftward shift in the ability of WAY-163909 to block apomorphine-induced climbing behavior, without any affect on apomorphine-induced stereotypy or an increased propensity for catalepsy. In the rat-conditioned avoidance model, WAY-163909 was combined with either haloperidol or clozapine at doses that individually produced reductions in avoidance response on the order of 10%, while the combination of WAY-163909 and either of the antipsychotics resulted in a greater than 70% reduction in avoidance, with no evidence of response failures, or pharmacokinetic interaction.ConclusionDoses of either haloperidol or clozapine, that failed to antagonize an MK-801 induced deficit in prepulse inhibition, significantly attenuated the sensory gating deficit when combined with WAY-163909. Data support the notion that 5-HT2C receptor agonists, co-administered with other marketed antipsychotics, allow for dose sparing with a more favorable side-effect profile.