Emma S. J. Robinson

Power failure: why small sample size undermines the reliability of neuroscience

A study with low statistical power has a reduced chance of detecting a true effect, but it is less well appreciated that low power also reduces the likelihood that a statistically significant result reflects a true effect. Here, we show that the average statistical power of studies in the neurosciences is very low. The consequences of this include overestimates of effect size and low reproducibility of results. There are also ethical dimensions to this problem, as unreliable research is inefficient and wasteful. Improving reproducibility in neuroscience is a key priority and requires attention to well-established but often ignored methodological principles.

Similar Effects of the Selective Noradrenaline Reuptake Inhibitor Atomoxetine on Three Distinct Forms of Impulsivity in the Rat

Atomoxetine is a noradrenaline-specific reuptake inhibitor used clinically for the treatment of childhood and adult attention deficit hyperactivity disorder (ADHD). Studies in human volunteers and patient groups have shown that atomoxetine improves stop-signal reaction time (SSRT) performance, an effect consistent with a reduction in motor impulsivity. However, ADHD is a heterogeneous disorder and it is of interest to determine whether atomoxetine is similarly effective against other forms of impulsivity, as well as the attentional impairment present in certain subtypes of ADHD. The present study examined the effects of atomoxetine on impulsivity using an analogous SSRT task in rats and two additional tests of impulsivity; delay discounting of reward and the five-choice serial reaction time task (5CSRTT), the latter providing an added assessment of sustained visual attention. Atomoxetine produced a significant dose-dependent speeding of SSRT. In addition, atomoxetine produced a selective, dose-dependent decrease in premature responding on the 5CSRTT. Finally, on the delay-discounting task, atomoxetine significantly decreased impulsivity by increasing preference for the large-value reward across increasing delay. These findings conclusively demonstrate that atomoxetine decreases several distinct forms of impulsivity in rats. The apparent contrast of these effects with stimulant drugs such as amphetamine and methylphenidate, which generally act to increase impulsivity on the 5CSRTT, may provide new insights into the mechanisms of action of stimulant and nonstimulant drugs in ADHD.

Opposing roles for 5-HT2A and 5-HT2C receptors in the nucleus accumbens on inhibitory response control in the 5-choice serial reaction time task.

Serotonin (5-HT) is thought to play an important role in the regulation of behavioral inhibition. Studies manipulating 5-HT function in the rodent brain indicate that 5-HT receptors regulate distinct forms of impulsive behavior, including impulsive responding in the 5-choice serial reaction time task (5CSRTT). The present study investigates the loci of effects mediated by 5-HT2A and 5-HT2C receptors in attention and inhibitory response control using microinfusions targeted at the nucleus accumbens (NAc), prelimbic cortex (PL) and infralimbic cortex (IL). Rats were implanted with bilateral guide cannulas and received infusions of the selective 5-HT2A receptor antagonist M100907 (0.1 and 0.3 μg) or selective 5-HT2C receptor antagonist SB242084 (0.1 and 0.5 μg) immediately prior to testing. The results show that intra-NAc infusions of M100907 significantly decrease impulsive responding on the 5CSRTT and at the highest dose increased omissions as well. By contrast, infusions of SB242084 into the NAc selectively and dose-dependently increased impulsivity. Neither M100907 nor SB242084 significantly altered impulsive responding following either intra-PL or intra-IL administration. However, SB242084 significantly decreased omissions following intra-PL administration (0.5 μg only). These data reveal opposing effects on impulsivity following 5-HT2A and 5-HT2C blockade in the NAc. Our results suggest that the NAc, but not the PL or IL, is implicated in the mediation of the effects of M100907 and SB242084 on inhibitory response control during baseline 5CSRTT performance.

Contrasting effects of selective lesions of nucleus accumbens core or shell on inhibitory control and amphetamine-induced impulsive behaviour

The core and shell subregions of the nucleus accumbens receive differential projections from areas of the medial prefrontal cortex that have dissociable effects on impulsive and perseverative responding. The contributions of these subregions to simple instrumental behaviour, inhibitory control and behavioural flexibility were investigated using a ‘forced choice’ task, various parameter manipulations and an omission schedule version of the task. Post‐training, selective core lesions were achieved with microinjections of quinolinic acid and shell lesions with ibotenic acid. After a series of behavioural task manipulations, rats were re‐stabilized on the standard version of the task and challenged with increasing doses of d‐amphetamine (vehicle, 0.5 or 1.0 mg/kg i.p. 30 min prior to test). Neither core‐ nor shell‐lesioned rats exhibited persistent deficits in simple instrumental behaviour or challenges to behavioural flexibility or inhibitory control. Significant differences between lesion groups were unmasked by d‐amphetamine challenge in the standard version of the forced task. Core lesions potentiated and shell lesions attenuated the dose‐dependent effect of d‐amphetamine on increasing anticipatory responses seen in sham rats. These data imply that the accumbens core and shell subregions do not play major roles in highly‐trained task performance or in challenges to behavioural control, but may have opposed effects following d‐amphetamine treatment. Specifically, they suggest the shell subregion to be necessary for dopaminergic activation driving amphetamine‐induced impulsive behaviour and the core subregion for the normal control of this behaviour via conditioned influences.

Modelling cognitive affective biases in major depressive disorder using rodents.

Major depressive disorder (MDD) affects more than 10% of the population, although our understanding of the underlying aetiology of the disease and how antidepressant drugs act to remediate symptoms is limited. Major obstacles include the lack of availability of good animal models that replicate aspects of the phenotype and tests to assay depression‐like behaviour in non‐human species. To date, research in rodents has been dominated by two types of assays designed to test for depression‐like behaviour: behavioural despair tests, such as the forced swim test, and measures of anhedonia, such as the sucrose preference test. These tests have shown relatively good predictive validity in terms of antidepressant efficacy, but have limited translational validity. Recent developments in clinical research have revealed that cognitive affective biases (CABs) are a key feature of MDD. Through the development of neuropsychological tests to provide objective measures of CAB in humans, we have the opportunity to use ‘reverse translation’ to develop and evaluate whether similar methods are suitable for research into MDD using animals. The first example of this approach was reported in 2004 where rodents in a putative negative affective state were shown to exhibit pessimistic choices in a judgement bias task. Subsequent work in both judgement bias tests and a novel affective bias task suggest that these types of assay may provide translational methods for studying MDD using animals. This review considers recent work in this area and the pharmacological and translational validity of these new animal models of CABs.

Endogenous β‐Carbolines as Clonidine‐Displacing Substances

Abstract: Endogenous β‐carbolines, such as harmane, are known to occur in mammalian species including humans. Radioligand binding studies have revealed that certain β‐carbolines display high affinity for both I1 and I2 imidazoline‐binding sites (IBS). Functional studies have shown that the β‐carboline harmane elicits many characteristics expected of an endogenous ligand IBS. This article discusses the evidence relating to β‐carbolines as endogenous ligands and presents a case for harmane and related compounds as endogenous ligands for IBS.

A Translational Rodent Assay of Affective Biases in Depression and Antidepressant Therapy

The subjective measures used to study mood disorders in humans cannot be replicated in animals; however, the increasing application of objective neuropsychological methods provides opportunities to develop translational animal tasks. Here we describe a novel behavioral approach, which has enabled us to investigate similar affective biases in rodents. In our affective bias test (ABT), rats encounter two independent positive experiences—the association between food reward and specific digging substrate—during discrimination learning sessions. These are performed on separate days under either neutral conditions or during a pharmacological or affective state manipulation. Affective bias is then quantified using a preference test where both previously rewarded substrates are presented together and the rat’s choices recorded. The absolute value of the experience is kept consistent and all other factors are counterbalanced so that any bias at recall can be attributed to treatment. Replicating previous findings from studies in healthy volunteers, we observe significant positive affective biases following acute treatment with typical (fluoxetine, citalopram, reboxetine, venlafaxine, clomipramine) and atypical antidepressants (agomelatine, mirtazapine), and significant negative affective biases following treatment with drugs associated with inducing negative affective states in humans (FG7142, rimonabant, 13-cis retinoic acid). We also observed that acute psychosocial stress and environmental enrichment induce significant negative and positive affective biases, respectively, and provide evidence that these affective biases involve memory consolidation. The positive and negative affective biases induced in our test also mirror the antidepressant and pro-depressant effects of these drugs in patients suggesting our test has both translational and predictive validity. Our results suggest that cognitive affective biases could contribute to drug- or stress-induced mood changes in people and support the hypothesis that a cognitive neuropsychological mechanism contributes to antidepressant drug efficacy.

In vitro and ex vivo distribution of [3H]harmane, an endogenous β-carboline, in rat brain

The endogenous beta-carboline, harmane, has been shown to bind to monoamine oxidase A (MAO-A) and a separate, high affinity, non-MAO site. Research in our laboratory has shown that harmane is an active component of clonidine-displacing substance (CDS), the proposed endogenous ligand for imidazoline binding sites (IBS). In the present study we have investigated the distribution of [3H]harmane in rat brain, and related the binding profile to the distribution of the MAO-A selective ligand [3H]Ro41-1049 and the I2BS ligand [3H]2-BFI. The in vivo distribution of [3H]harmane following intravenous administration was also investigated. Receptor autoradiography revealed a highly significant correlation for the distribution of [3H]harmane and [3H]Ro41-1049, and a significant correlation for [3H]harmane and the I2BS ligand [3H]2-BFI. The in vivo distribution of [3H]harmane suggests that the ligand accumulates in the adrenal gland and throughout the brain with the primary route of excretion occurring via the duodenum. In conclusion, these studies have shown that [3H]harmane labels a population of binding sites that reflect the distribution of MAO-A. Further evidence for a non-MAO, IBS [3H]harmane population has not been shown but the high level of expression of the MAO-A site is likely to have masked the much smaller population of I2BS.

Study of Novel Selective mGlu2 Agonist in the Temporo-Ammonic Input to CA1 Neurons Reveals Reduced mGlu2 Receptor Expression in a Wistar Substrain with an Anxiety-Like Phenotype

Group II metabotropic receptors (mGluRs) regulate central synaptic transmission by modulating neurotransmitter release. However, the lack of pharmacological tools differentiating between mGlu2 and mGlu3 receptors has hampered identification of the roles of these two receptor subtypes. We have used LY395756 [(1SR,2SR,4RS,5RS,6SR)-2-amino-4-methylbicyclo[3.1.0]-hexane2,6-dicarboxylic], an agonist at mGlu2 receptors and an antagonist at mGlu3 receptors in cell lines, to investigate the roles of these receptors in the temporo-ammonic path from entorhinal cortex to CA1–stratum lacunosum moleculare in rat hippocampal slices. Surprisingly, the degree of inhibition of the field EPSP induced by LY395756 fell into two distinct groups, with EC50 values of <1 μm and >100 μm. In “sensitive” slices, LY395756 had additive actions with a mixed mGlu2/mGlu3 agonist, DCG-IV [(2S,2′R,3′R)-2-(2′,3′-dicarboxycyclopropyl)glycine], whereas in “insensitive” slices, LY395756 reduced the effect of DCG-IV, with an IC50 of ∼1 μm. This separation into sensitive and insensitive slices could be explained by LY395756 acting as an mGlu2 agonist and mGlu3 antagonist, respectively, a finding supported by data from mice lacking these receptors. The heterogeneity was correlated with differences in expression levels of mGlu2 receptors within our Wistar colony and other Wistar substrains. The initial search for a behavioral correlate indicated that rats lacking mGlu2 receptors showed anxiety-like behavior in open-field and elevated plus maze assays. These findings have implications for rat models of psychiatric disease and are especially pertinent given that mGlu2 receptors are targets for compounds under development for anxiety.

Evaluation of a novel translational task for assessing emotional biases in different species

Changes in the processing of emotional information are key features of affective disorders. Neuropsychological tests based on emotional faces or words are used to detect emotional/affective biases in humans, but these tests are not applicable to animal species. In the present study, we investigated whether a novel affective tone discrimination task (ATDT), developed to study emotion-related behaviour in rats, could also be used to quantify changes in affective states in humans. To date, the methods used in human neuropsychology have not been applicable to animal experiments. Participants completed a training session in which they learnt to discriminate specific tone frequencies and to correctly respond in order to gain emotionally valenced outcomes, to obtain rewards (money), or to avoid punishment (an aversive sound clip). During a subsequent test session, additional ambiguous probe tones were presented at frequencies intermediate between the reward and avoidance paired tones. At the end of the task, participants completed self-report questionnaires. All participants made more avoidance responses to the most ambiguous tone cues, suggesting a bias towards avoidance of punishment. Individual differences in the degrees of bias observed were correlated with anxiety measures, suggesting the task’s sensitivity to differences in state anxiety within a healthy population. Further studies in clinical populations will be necessary to assess the task’s sensitivity to pathological anxiety states. These data suggest that this affective tone discrimination task provides a novel method to study cognitive affective biases in different species, including humans, and offers a novel assessment to study anxiety.