Preclinical screening for antidepressant activity - shifting focus away from the Forced Swim Test to the use of translational biomarkers.

Depression is the world's predominant mental health problem and a leading cause of disability. Neuropharmacological research has not yet advanced treatments to sufficiently meet clinical need, largely due to the failure of animal models to predict clinical efficacy. The forced swim test (FST) has been extensively used in the field of antidepressant research but has been under scrutiny due to its perceived severity to animals. Any use of animals in experiments and testing must have a scientific or regulatory purpose and researchers need to ensure that there is no scientifically valid alternative. However, regulatory requirements have been incorrectly cited as a reason to support the use of the FST. More research is required on tests that do not involve stressing animals as replacements for the FST. Non-behavioural neurochemical measures might provide a means to advance neuropharmacological developments while reducing animal suffering. For example, brain-derived neurotrophic factor (BDNF) may be promising.

Assessing the Cognitive Translational Potential of a Mouse Model of the 22q11.2 Microdeletion Syndrome.

A chromosomal microdeletion at the 22q11.2 locus is associated with extensive cognitive impairments, schizophrenia and other psychopathology in humans. Previous reports indicate that mouse models of the 22q11.2 microdeletion syndrome (22q11.2DS) may model the genetic basis of cognitive deficits relevant for neuropsychiatric disorders such as schizophrenia. To assess the models usefulness for drug discovery, a novel mouse (Df(h22q11)/+) was assessed in an extensive battery of cognitive assays by partners within the NEWMEDS collaboration (Innovative Medicines Initiative Grant Agreement No. 115008). This battery included classic and touchscreen-based paradigms with recognized sensitivity and multiple attempts at reproducing previously published findings in 22q11.2DS mouse models. This work represents one of the most comprehensive reports of cognitive functioning in a transgenic animal model. In accordance with previous reports, there were non-significant trends or marginal impairment in some tasks. However, the Df(h22q11)/+ mouse did not show comprehensive deficits; no robust impairment was observed following more than 17 experiments and 14 behavioral paradigms. Thus - within the current protocols - the 22q11.2DS mouse model fails to mimic the cognitive alterations observed in human 22q11.2 deletion carriers. We suggest that the 22q11.2DS model may induce liability for cognitive dysfunction with additional "hits" being required for phenotypic expression.

Task-induced modulation of intrinsic functional connectivity networks in the behaving rat.

While resting-state functional magnetic resonance imaging can probe intrinsic network connectivity in both human and rodent brain, behavioral modulation of these connectivity patterns has not yet been demonstrated in the rodent due to the requirements of immobilization or anesthesia for MRI scanning. To enable the effects of behavioral tasks on functional connectivity to be measured in freely moving, awake rats, implanted carbon paste electrodes (CPEs) were used to monitor low-frequency fluctuations of tissue oxygenation. Rats were implanted with CPEs in two nodes of the default mode network (DMN) and two nodes in a lateral cortical network, revealing amperometric oxygen correlation patterns consistent with imaging studies. Using a block design study where rats alternated between sustained periods of instrumental response and unscheduled spontaneous behavior, task-induced decreases in functional connectivity were observed between the DMN node pair, but not in the distinct lateral cortical network, demonstrating network-specific modulation of functional connectivity.

Differential contributions of infralimbic prefrontal cortex and nucleus accumbens during reward-based learning and extinction.

Using environmental cues for the prediction of future events is essential for survival. Such cue-outcome associations are thought to depend on mesolimbic circuitry involving the nucleus accumbens (NAc) and prefrontal cortex (PFC). Several studies have identified roles for both NAc and PFC in the expression of stable goal-directed behaviors, but much remains unknown about their roles during learning of such behaviors. To further address this question, we used in vivo oxygen amperometry, a proxy for blood oxygen level-dependent (BOLD) signal measurement in human functional magnetic resonance imaging, in rats performing a cued lever-pressing task requiring discrimination between a rewarded and nonrewarded cue. Simultaneous oxygen recordings were obtained from infralimbic PFC (IFC) and NAc throughout both acquisition and extinction of this task. Activation of NAc was specifically observed following rewarded cue onset during the entire acquisition phase and also during the first days of extinction. In contrast, IFC activated only during the earliest periods of acquisition and extinction, more specifically to the nonrewarded cue. Thus, in vivo oxygen amperometry permits a novel, stable form of longitudinal analysis of brain activity in behaving animals, allowing dissociation of the roles of different brain regions over time during learning of reward-driven instrumental action. The present results offer a unique temporal perspective on how NAc may promote actions directed toward anticipated positive outcome throughout learning, while IFC might suppress actions that no longer result in reward, but only during critical periods of learning.

A targeted multiplexed proteomic investigation identifies ketamine-induced changes in immune markers in rat serum and expression changes in protein kinases/phosphatases in rat brain.

There is substantial interest in the N-methyl-d-aspartate (NMDA) receptor antagonist ketamine in psychiatric research because it exerts acute psychotomimetic and rapid antidepressant effects in rodents and humans. Here, we investigated proteomic changes in brain and serum after acute treatment of rats with ketamine using two targeted proteomic profiling methods. Multiplex immunoassay profiling of serum identified altered levels of interleukin 4, tumor necrosis factor alpha, and fibroblast growth factor 9, suggesting a link between ketamine exposure and peripheral inflammation and growth factor dysregulation. Selected reaction monitoring mass spectrometry profiling of rat brain tissue found that proteomic changes occurred in the frontal cortex and to a greater extent in the hippocampus. This involved changes in signaling kinases and proteases such as protein kinase C beta, neurochondrin (NCDN), calcineurin, extracellular signal-regulated kinsase 1 (ERK1), and mammalian target of rapamycin (MTOR). Furthermore, altered levels were found for proteins associated with neurotransmitter metabolism (mitochondrial aspartate aminotransferase, catechol O-methyl transferase, synaptic vesicle endo-/exocytosis (vesicle fusing ATPase (NSF), synapsin 1 (SYN1), syndapin-1 (PACN1)). Consistent with previous global proteomic studies, we confirmed known changes in mitochondrial complex I, prohibitin (PHB) and neurofilament proteins (neurofilament light chain and α-internexin (AINX)). Taken together, the proteomic changes parallel those described in human psychiatric pathology. The results will help to elucidate ketamine's mechanism of action, which will facilitate development of novel drugs for the treatment of schizophrenia and major depressive disorder.

Prefrontal cholinergic mechanisms instigating shifts from monitoring for cues to cue-guided performance: converging electrochemical and fMRI evidence from rats and humans.

We previously reported involvement of right prefrontal cholinergic activity in veridical signal detection. Here, we first recorded real-time acetylcholine release in prefrontal cortex (PFC) during specific trial sequences in rats performing a task requiring signal detection as well as rejection of nonsignal events. Cholinergic release events recorded with subsecond resolution ("transients") were observed only during signal-hit trials, not during signal-miss trials or nonsignal events. Moreover, cholinergic transients were not observed for consecutive hits; instead they were limited to signal-hit trials that were preceded by factual or perceived nonsignal events ("incongruent hits"). This finding suggests that these transients mediate shifts from a state of perceptual attention, or monitoring for cues, to cue-evoked activation of response rules and the generation of a cue-directed response. Next, to determine the translational significance of the cognitive operations supporting incongruent hits we used a version of the task previously validated for use in research in humans and blood oxygenation level-dependent (BOLD)-functional magnetic resonance imaging. Incongruent hits activated a region in the right rostral PFC (Brodmann area 10). Furthermore, greater prefrontal activation was correlated with faster response times for incongruent hits. Finally, we measured tissue oxygen in rats, as a proxy for BOLD, and found prefrontal increases in oxygen levels solely during incongruent hits. These cross-species studies link a cholinergic response to a prefrontal BOLD activation and indicate that these interrelated mechanisms mediate the integration of external cues with internal representations to initiate and guide behavior.

Characterisation of a microelectrochemical biosensor for real-time detection of brain extracellular d-serine.

A modified development protocol and concomitant characterisation of a first generation biosensor for the detection of brain extracellular d-serine is reported. Functional parameters important for neurochemical monitoring, including sensor sensitivity, O2 interference, selectivity, shelf-life and biocompatibility were examined. Construction and development involved the enzyme d-amino acid oxidase (DAAO), utilising a dip-coating immobilisation method employing a new extended drying approach. The resultant Pt-based polymer enzyme composite sensor achieved high sensitivity to d-serine (0.76 ± 0.04 nA mm-2. µM-1) and a low µM limit of detection (0.33 ± 0.02 µM). The in-vitro response time was within the solution stirring time, suggesting potential sub-second in-vivo response characteristics. Oxygen interference studies demonstrated a 1 % reduction in current at 50 µM O2 when compared to atmospheric O2 levels (200 µM), indicating that the sensor can be used for reliable neurochemical monitoring of d-serine, free from changes in current associated with physiological O2 fluctuations. Potential interference signals generated by the principal electroactive analytes present in the brain were minimised by using a permselective layer of poly(o-phenylenediamine), and although several d-amino acids are possible substrates for DAAO, their physiologically relevant signals were small relative to that for d-serine. Additionally, changing both temperature and pH over possible in vivo ranges (34-40 °C and 7.2-7.6 respectively) resulted in no significant effect on performance. Finally, the biosensor was implanted in the striatum of freely moving rats and used to monitor physiological changes in d-serine over a two-week period.

Hemodynamic responses in amygdala and hippocampus distinguish between aversive and neutral cues during Pavlovian fear conditioning in behaving rats.

Lesion and electrophysiological studies in rodents have identified the amygdala and hippocampus (HPC) as key structures for Pavlovian fear conditioning, but human functional neuroimaging studies have not consistently found activation of these structures. This could be because hemodynamic responses cannot detect the sparse neuronal activity proposed to underlie conditioned fear. Alternatively, differences in experimental design or fear levels could account for the discrepant findings between rodents and humans. To help distinguish between these alternatives, we used tissue oxygen amperometry to record hemodynamic responses from the basolateral amygdala (BLA), dorsal HPC (dHPC) and ventral HPC (vHPC) in freely-moving rats during the acquisition and extinction of conditioned fear. To enable specific comparison with human studies we used a discriminative paradigm, with one auditory cue [conditioned stimulus (CS)+] that was always followed by footshock, and another auditory cue (CS-) that was never followed by footshock. BLA tissue oxygen signals were significantly higher during CS+ than CS- trials during training and early extinction. In contrast, they were lower during CS+ than CS- trials by the end of extinction. dHPC and vHPC tissue oxygen signals were significantly lower during CS+ than CS- trials throughout extinction. Thus, hemodynamic signals in the amygdala and HPC can detect the different patterns of neuronal activity evoked by threatening vs. neutral stimuli during fear conditioning. Discrepant neuroimaging findings may be due to differences in experimental design and/or fear levels evoked in participants. Our methodology offers a way to improve translation between rodent models and human neuroimaging.

Distinct hippocampal-prefrontal neural assemblies coordinate memory encoding, maintenance, and recall.

Short-term memory enables incorporation of recent experience into subsequent decision-making. This processing recruits both the prefrontal cortex and hippocampus, where neurons encode task cues, rules, and outcomes. However, precisely which information is carried when, and by which neurons, remains unclear. Using population decoding of activity in rat medial prefrontal cortex (mPFC) and dorsal hippocampal CA1, we confirm that mPFC populations lead in maintaining sample information across delays of an operant non-match to sample task, despite individual neurons firing only transiently. During sample encoding, distinct mPFC subpopulations joined distributed CA1-mPFC cell assemblies hallmarked by 4-5 Hz rhythmic modulation; CA1-mPFC assemblies re-emerged during choice episodes but were not 4-5 Hz modulated. Delay-dependent errors arose when attenuated rhythmic assembly activity heralded collapse of sustained mPFC encoding. Our results map component processes of memory-guided decisions onto heterogeneous CA1-mPFC subpopulations and the dynamics of physiologically distinct, distributed cell assemblies.

Improving Translational Relevance in Preclinical Psychopharmacology (iTRIPP).

Animal models are important in preclinical psychopharmacology to study mechanisms and potential treatments for psychiatric disorders. A working group of 14 volunteers, comprising an international team of researchers from academia and industry, convened in 2021 to discuss how to improve the translational relevance and interpretation of findings from animal models that are used in preclinical psychopharmacology. The following paper distils the outcomes of the working group's discussions into 10 key considerations for the planning and reporting of behavioural studies in animal models relevant to psychiatric disorders. These form the iTRIPP guidelines (Improving Translational Relevance In Preclinical Psychopharmacology). These guidelines reflect the key considerations that the group thinks will likely have substantial impact in terms of improving the translational relevance of behavioural studies in animal models that are used to study psychiatric disorders and their treatment. They are relevant to the research community when drafting and reviewing manuscripts, presentations and grant applications. The iTRIPP guidelines are intended to complement general recommendations for planning and reporting animal studies that have been published elsewhere, by enabling researchers to fully consider the most appropriate animal model for the research purpose and to interpret their findings appropriately. This in turn will increase the clinical benefit of such research and is therefore important not only for the scientific community but also for patients and the lay public.

Dysfunctional prefrontal cortical network activity and interactions following cannabinoid receptor activation.

Coordinated activity spanning anatomically distributed neuronal networks underpins cognition and mediates limbic-cortical interactions during learning, memory, and decision-making. We used CP55940, a potent agonist of brain cannabinoid receptors known to disrupt coordinated activity in hippocampus, to investigate the roles of network oscillations during hippocampal and medial prefrontal cortical (mPFC) interactions in rats. During quiet wakefulness and rest, CP55940 dose-dependently reduced 0.1-30 Hz local field potential power in CA1 of the hippocampus while concurrently decreasing 30-100 Hz power in mPFC; these contrasting population-level effects were paralleled by differential effects on underlying single-unit activity in the two structures. During decision-making phases of a spatial working memory task, CP5540-induced deficits in hippocampal theta and prefrontal gamma oscillations were observed alongside disrupted theta-frequency coherence between the two structures. These changes in coordinated limbic-cortical network activities correlated with (1) reduced accuracy of task performance, (2) impaired phase-locking of prefrontal single-unit spiking to the local gamma and hippocampal theta rhythms, and (3) impaired task-dependent activity in a subset of mPFC units. In addition to highlighting the importance of CA1-mPFC network oscillations for cognition, these results implicate disrupted theta-frequency coordination of CA1-mPFC activity in the cognitive deficits caused by exogenous activation of brain cannabinoid receptors.

Changes in reward-related signals in the rat nucleus accumbens measured by in vivo oxygen amperometry are consistent with fMRI BOLD responses in man.

Real-time in vivo oxygen amperometry, a technique that allows measurement of regional brain tissue oxygen (O(2)) has been previously shown to bear relationship to the BOLD signal measured with functional magnetic resonance imaging (fMRI) protocols. In the present study, O(2) amperometry was applied to the study of reward processing in the rat nucleus accumbens to validate the technique with a behavioural process known to cause robust signals in human neuroimaging studies. After acquisition of a cued-lever pressing task a robust increase in O(2) tissue levels was observed in the nucleus accumbens specifically following a correct lever press to the rewarded cue. This O(2) signal was modulated by cue reversal but not lever reversal, by differences in reward magnitudes and by the motivational state of the animal consistent with previous reports of the role of the nucleus accumbens in both the anticipation and representation of reward value. Moreover, this modulation by reward value was related more to the expected incentive value rather than the hedonic value of reward, also consistent with previous reports of accumbens coding of "wanting" of reward. Altogether, these results show striking similarities to those obtained in human fMRI studies suggesting the use of oxygen amperometry as a valid surrogate for fMRI in animals performing cognitive tasks, and a powerful approach to bridge between different techniques of measurement of brain function.

Close temporal coupling of neuronal activity and tissue oxygen responses in rodent whisker barrel cortex.

Neuronal activity elicits metabolic and vascular responses, during which oxygen is first consumed and then supplied to the tissue via an increase in cerebral blood flow. Understanding the spatial and temporal dynamics of blood and tissue oxygen (To2) responses following neuronal activity is crucial for understanding the physiological basis of functional neuroimaging signals. However, our knowledge is limited because previous To2 measurements have been made at low temporal resolution (>100 ms). Here we recorded To2 at high temporal resolution (1 ms), simultaneously with co-localized field potentials, at several cortical depths from the whisker region of the somatosensory cortex in anaesthetized rats and mice. Stimulation of the whiskers produced rapid, laminar-specific changes in To2. Positive To2 responses (i.e. increases) were observed in the superficial layers within 50 ms of stimulus onset, faster than previously reported. Negative To2 responses (i.e. decreases) were observed in the deeper layers, with maximal amplitude in layer IV, within 40 ms of stimulus onset. The amplitude of the negative, but not the positive, To2 response correlated with local field potential amplitude. Disruption of neurovascular coupling, via nitric oxide synthase inhibition, abolished positive To2 responses to whisker stimulation in the superficial layers and increased negative To2 responses in all layers. Our data show that To2 responses occur rapidly following neuronal activity and are laminar dependent.

Time to re-engage psychiatric drug discovery by strengthening confidence in preclinical psychopharmacology.

BACKGROUND: There is urgent need for new medications for psychiatric disorders. Mental illness is expected to become the leading cause of disability worldwide by 2030. Yet, the last two decades have seen the pharmaceutical industry withdraw from psychiatric drug discovery after costly late-stage trial failures in which clinical efficacy predicted pre-clinically has not materialised, leading to a crisis in confidence in preclinical psychopharmacology. METHODS: Based on a review of the relevant literature, we formulated some principles for improving investment in translational neuroscience aimed at psychiatric drug discovery. RESULTS: We propose the following 8 principles that could be used, in various combinations, to enhance CNS drug discovery: (1) consider incorporating the NIMH Research Domain Criteria (RDoC) approach; (2) engage the power of translational and systems neuroscience approaches; (3) use disease-relevant experimental perturbations; (4) identify molecular targets via genomic analysis and patient-derived pluripotent stem cells; (5) embrace holistic neuroscience: a partnership with psychoneuroimmunology; (6) use translational measures of neuronal activation; (7) validate the reproducibility of findings by independent collaboration; and (8) learn and reflect. We provide recent examples of promising animal-to-human translation of drug discovery projects and highlight some that present re-purposing opportunities. CONCLUSIONS: We hope that this review will re-awaken the pharma industry and mental health advocates to the opportunities for improving psychiatric pharmacotherapy and so restore confidence and justify re-investment in the field.

Real-time electrochemical monitoring of brain tissue oxygen: a surrogate for functional magnetic resonance imaging in rodents.

Long-term in-vivo electrochemistry (LIVE) enables real-time monitoring and measurement of brain metabolites. In this study we have simultaneously obtained blood oxygenation level dependent (BOLD) fMRI and amperometric tissue O(2) data from rat cerebral cortex, during both increases and decreases in inspired O(2) content. BOLD and tissue O(2) measurements demonstrated close correlation (r=0.7898) during complete (0%) O(2) removal, with marked negative responses occurring ca. 30s after the onset of O(2) removal. Conversely, when the inspired O(2) was increased (50, 70 and 100% O(2) for 1min) similar positive rapid changes (ca. 15s) in both the BOLD and tissue O(2) signals were observed. These findings demonstrate, for the first time, the practical feasibility of obtaining real-time metabolite information during fMRI acquisition, and that tissue O(2) concentration monitored using an O(2) sensor can serve as an index of changes in the magnitude of the BOLD response. As LIVE O(2) sensors can be used in awake animals performing specific behavioural tasks the technique provides a viable animal surrogate of human fMRI experimentation.

Amphetamine disrupts haemodynamic correlates of prediction errors in nucleus accumbens and orbitofrontal cortex.

In an uncertain world, the ability to predict and update the relationships between environmental cues and outcomes is a fundamental element of adaptive behaviour. This type of learning is typically thought to depend on prediction error, the difference between expected and experienced events and in the reward domain that has been closely linked to mesolimbic dopamine. There is also increasing behavioural and neuroimaging evidence that disruption to this process may be a cross-diagnostic feature of several neuropsychiatric and neurological disorders in which dopamine is dysregulated. However, the precise relationship between haemodynamic measures, dopamine and reward-guided learning remains unclear. To help address this issue, we used a translational technique, oxygen amperometry, to record haemodynamic signals in the nucleus accumbens (NAc) and orbitofrontal cortex (OFC), while freely moving rats performed a probabilistic Pavlovian learning task. Using a model-based analysis approach to account for individual variations in learning, we found that the oxygen signal in the NAc correlated with a reward prediction error, whereas in the OFC it correlated with an unsigned prediction error or salience signal. Furthermore, an acute dose of amphetamine, creating a hyperdopaminergic state, disrupted rats' ability to discriminate between cues associated with either a high or a low probability of reward and concomitantly corrupted prediction error signalling. These results demonstrate parallel but distinct prediction error signals in NAc and OFC during learning, both of which are affected by psychostimulant administration. Furthermore, they establish the viability of tracking and manipulating haemodynamic signatures of reward-guided learning observed in human fMRI studies by using a proxy signal for BOLD in a freely behaving rodent.

AMPA receptor potentiation can prevent ethanol-induced intoxication.

We present a substantial series of behavioral and imaging experiments, which demonstrate, for the first time, that increasing AMPA receptor-mediated neurotransmission via administration of potent and selective biarylsulfonamide AMPA potentiators LY404187 and LY451395 reverses the central effects of an acutely intoxicating dose of ethanol in the rat. Using pharmacological magnetic resonance imaging (phMRI), we observed that LY404187 attenuated ethanol-induced reductions in blood oxygenation level dependent (BOLD) in the anesthetized rat brain. A similar attenuation was apparent when measuring local cerebral glucose utilization (LCGU) via C14-2-deoxyglucose autoradiography in freely moving conscious rats. Both LY404187 and LY451395 significantly and dose-dependently reversed ethanol-induced deficits in both motor coordination and disruptions in an operant task where animals were trained to press a lever for food reward. Both prophylactic and acute intervention treatment with LY404187 reversed ethanol-induced deficits in motor coordination. Given that LY451395 and related AMPA receptor potentiators/ampakines are tolerated in both healthy volunteers and elderly patients, these data suggest that such compounds may form a potential management strategy for acute alcohol intoxication.

Ligands selective for alpha4beta2 but not alpha3beta4 or alpha7 nicotinic receptors generalise to the nicotine discriminative stimulus in the rat.

RATIONALE: Nicotine produces behavioural effects that are potentially related to its interaction with diverse nicotinic acetylcholine receptor populations. Evidence from gene deletion studies suggests that the interoceptive stimulus properties of nicotine are mediated by heteromeric high-affinity receptors containing alpha4beta2 subunits. Mice lacking beta2 subunits do not discriminate nicotine (Shoaib et al., Neuropharmacology, 42:530-539, 2002), and nicotine does not elicit dopamine release in these animals (Grady et al., J Neurochem, 76:258-268, 2001). The stimulus properties of nicotine can be detected in rats using a two-lever operant drug discrimination paradigm, allowing them to be classified pharmacologically using ligands with selectivity for receptors containing alpha4beta2, alpha3beta4 or alpha7 subunits. MATERIALS AND METHODS: Rats trained to discriminate 0.4 mg/kg nicotine from vehicle were given the nicotinic receptor agonists, cytisine, varenicline, TC2559, ABT-594, A-85380 (all having high affinity but varying selectivity for alpha4beta2-containing receptors), and WO 03/062224 and WO 01/60821A1 (selective for beta4- and alpha7-containing receptors, respectively). In separate studies, WO 03/062224 was used as the training stimulus. RESULTS: Nicotine, TC-2559, A-85380 and ABT-594 showed dose-dependent and complete stimulus substitution, whilst WO 03/062224 and WO 01/60821A1 were completely without effect. Cytisine and varenicline showed partial generalisation, consistent with their partial agonist activity at nicotinic receptors eliciting dopamine release in rat striatal slices. After almost 50 training sessions with WO 03/062224, there was no clear evidence that an alpha3beta4 receptor agonist could sustain a discriminable stimulus. CONCLUSION: Substitution to the nicotine discriminative stimulus required high-affinity and high intrinsic activity at beta2 but not at beta4- or at alpha7-containing nicotinic receptors.

TaiNi: Maximizing research output whilst improving animals' welfare in neurophysiology experiments.

Understanding brain function at the cell and circuit level requires representation of neuronal activity through multiple recording sites and at high sampling rates. Traditional tethered recording systems restrict movement and limit the environments suitable for testing, while existing wireless technology is still too heavy for extended recording in mice. Here we tested TaiNi, a novel ultra-lightweight (<2 g) low power wireless system allowing 72-hours of recording from 16 channels sampled at ~19.5 KHz (9.7 KHz bandwidth). We captured local field potentials and action-potentials while mice engaged in unrestricted behaviour in a variety of environments and while performing tasks. Data was synchronized to behaviour with sub-second precision. Comparisons with a state-of-the-art wireless system demonstrated a significant improvement in behaviour owing to reduced weight. Parallel recordings with a tethered system revealed similar spike detection and clustering. TaiNi represents a significant advance in both animal welfare in electrophysiological experiments, and the scope for continuously recording large amounts of data from small animals.

The IMPROVE Guidelines (Ischaemia Models: Procedural Refinements Of in Vivo Experiments).

Most in vivo models of ischaemic stroke target the middle cerebral artery and a spectrum of stroke severities, from mild to substantial, can be achieved. This review describes opportunities to improve the in vivo modelling of ischaemic stroke and animal welfare. It provides a number of recommendations to minimise the level of severity in the most common rodent models of middle cerebral artery occlusion, while sustaining or improving the scientific outcomes. The recommendations cover basic requirements pre-surgery, selecting the most appropriate anaesthetic and analgesic regimen, as well as intraoperative and post-operative care. The aim is to provide support for researchers and animal care staff to refine their procedures and practices, and implement small incremental changes to improve the welfare of the animals used and to answer the scientific question under investigation. All recommendations are recapitulated in a summary poster (see supplementary information).