A searchable image resource of Drosophila GAL4 driver expression patterns with single neuron resolution.

Precise, repeatable genetic access to specific neurons via GAL4/UAS and related methods is a key advantage of Drosophila neuroscience. Neuronal targeting is typically documented using light microscopy of full GAL4 expression patterns, which generally lack the single-cell resolution required for reliable cell type identification. Here, we use stochastic GAL4 labeling with the MultiColor FlpOut approach to generate cellular resolution confocal images at large scale. We are releasing aligned images of 74,000 such adult central nervous systems. An anticipated use of this resource is to bridge the gap between neurons identified by electron or light microscopy. Identifying individual neurons that make up each GAL4 expression pattern improves the prediction of split-GAL4 combinations targeting particular neurons. To this end, we have made the images searchable on the NeuronBridge website. We demonstrate the potential of NeuronBridge to rapidly and effectively identify neuron matches based on morphology across imaging modalities and datasets.

Acute D-serine treatment produces antidepressant-like effects in rodents.

Research suggests that dysfunctional glutamatergic signalling may contribute to depression, a debilitating mood disorder affecting millions of individuals worldwide. Ketamine, a N-methyl-D-aspartate (NMDA) receptor antagonist, exerts rapid antidepressant effects in approximately 70% of patients. Glutamate evokes the release of D-serine from astrocytes and neurons, which then acts as a co-agonist and binds at the glycine site on the NR1 subunit of NMDA receptors. Several studies have implicated glial deficits as one of the underlying facets of the neurobiology of depression. The present study tested the hypothesis that D-serine modulates behaviours related to depression. The behavioural effects of a single, acute D-serine administration were examined in several rodent tests of antidepressant-like effects, including the forced swim test (FST), the female urine sniffing test (FUST) following serotonin depletion, and the learned helplessness (LH) paradigm. D-serine significantly reduced immobility in the FST without affecting general motor function. Both D-serine and ketamine significantly rescued sexual reward-seeking deficits caused by serotonin depletion in the FUST. Finally, D-serine reversed LH behaviour, as measured by escape latency, number of escapes, and percentage of mice developing LH. Mice lacking NR1 expression in forebrain excitatory neurons exhibited a depression-like phenotype in the same behavioural tests, and did not respond to D-serine treatment. These findings suggest that D-serine produces antidepressant-like effects and support the notion of complex glutamatergic dysfunction in depression. It is unclear whether D-serine has a convergent influence on downstream synaptic plasticity cascades that may yield a similar therapeutic profile to NMDA antagonists like ketamine.

Animal models of suicide-trait-related behaviors.

Although antidepressants are moderately effective in treating major depressive disorder (MDD), concerns have arisen that selective serotonin-reuptake inhibitors (SSRIs) are associated with suicidal thinking and behavior, especially in children, adolescents and young adults. Almost no experimental research in model systems has considered the mechanisms by which SSRIs might be associated with this potential side effect in some susceptible individuals. Suicide is a complex behavior and impossible to fully reproduce in an animal model. However, by investigating traits that show strong cross-species parallels in addition to associations with suicide in humans, animal models might elucidate the mechanisms by which SSRIs are associated with suicidal thinking and behavior. Traits linked with suicide in humans that can be successfully modeled in rodents include aggression, impulsivity, irritability and hopelessness/helplessness. Modeling these relevant traits in animals can help to clarify the impact of SSRIs on these traits, suggesting avenues for reducing suicide risk in this vulnerable population.

An unbiased template of the Drosophila brain and ventral nerve cord.

The fruit fly Drosophila melanogaster is an important model organism for neuroscience with a wide array of genetic tools that enable the mapping of individual neurons and neural subtypes. Brain templates are essential for comparative biological studies because they enable analyzing many individuals in a common reference space. Several central brain templates exist for Drosophila, but every one is either biased, uses sub-optimal tissue preparation, is imaged at low resolution, or does not account for artifacts. No publicly available Drosophila ventral nerve cord template currently exists. In this work, we created high-resolution templates of the Drosophila brain and ventral nerve cord using the best-available technologies for imaging, artifact correction, stitching, and template construction using groupwise registration. We evaluated our central brain template against the four most competitive, publicly available brain templates and demonstrate that ours enables more accurate registration with fewer local deformations in shorter time.

Genome-wide gene expression profiling in GluR1 knockout mice: key role of the calcium signaling pathway in glutamatergically mediated hippocampal transmission.

Alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptors (AMPARs) convey fast synaptic transmission in the CNS and mediate various forms of hippocampal plasticity. Disruption of glutamate receptor type 1 (GluR1), a member of the AMPAR family, causes synaptic alterations and learning/memory deficits in mice. To gain mechanistic insight into the synaptic and behavioral changes associated with GluR1 deletion, hippocampal genome-wide expression profiling was conducted using groups of GluR1 knockout (KO) mice and their wild-type littermates. Regulation of 38 genes was found to be altered more than 30% (P < 0.01, n = 8), and seven of these genes were studied with additional quantitative experiments. A large portion of the altered genes encoded molecules involved in calcium signaling, including calcium channel components, calcium-binding proteins and calcium-calmodulin-dependent protein kinase II subunits. At the protein level, we further evaluated some genes in the calcium pathway that were altered in GluR1 KO mice. Protein levels of two key molecules in the calcium pathway - GluR, ionotropic, N-methyl-d-aspartate-1 and calcium/calmodulin-dependent protein kinase II alpha - showed similar changes to those observed in mRNA levels. These findings raise the possibility that calcium signaling and other plasticity molecules may contribute to the hippocampal plasticity and behavioral deficits observed in GluR1 KO mice.

Mapping Neurotransmitter Identity in the Whole-Mount Drosophila Brain Using Multiplex High-Throughput Fluorescence in Situ Hybridization.

Identifying the neurotransmitters used by specific neurons is a critical step in understanding the function of neural circuits. However, methods for the consistent and efficient detection of neurotransmitter markers remain limited. Fluorescence in situ hybridization (FISH) enables direct labeling of type-specific mRNA in neurons. Recent advances in FISH allow this technique to be carried out in intact tissue samples such as whole-mount Drosophila melanogaster brains. Here, we present a FISH platform for high-throughput detection of eight common neurotransmitter phenotypes in Drosophila brains. We greatly increase FISH throughput by processing samples mounted on coverslips and optimizing fluorophore choice for each probe to facilitate multiplexing. As application examples, we demonstrate cases of neurotransmitter coexpression, reveal neurotransmitter phenotypes of specific cell types, and explore the onset of neurotransmitter expression in the developing optic lobe. Beyond neurotransmitter markers, our protocols can in principle be used for large-scale FISH detection of any mRNA in whole-mount fly brains.

Optimization of fluorophores for chemical tagging and immunohistochemistry of Drosophila neurons.

The use of genetically encoded 'self-labeling tags' with chemical fluorophore ligands enables rapid labeling of specific cells in neural tissue. To improve the chemical tagging of neurons, we synthesized and evaluated new fluorophore ligands based on Cy, Janelia Fluor, Alexa Fluor, and ATTO dyes and tested these with recently improved Drosophila melanogaster transgenes. We found that tissue clearing and mounting in DPX substantially improves signal quality when combined with specific non-cyanine fluorophores. We compared and combined this labeling technique with standard immunohistochemistry in the Drosophila brain.

Gastric preloads of corn oil and mineral oil produce different patterns of increases of c-Fos-like immunoreacitve cells in the brain of 9-12 day-old rats.

Equivolumetric gastric preloads of corn oil and mineral oil administered to rats on postnatal day 12 (P12) inhibited intake equally during a 30-min test of independent ingestion (II), but preloads of corn oil inhibited intake significantly more than preloads of mineral oil on P15 and P18 [Weller, A., Gispan, I.H., Armony-Sivan, R., Ritter, R.C., Smith, G.P., 1997. Preloads of corn oil inhibit independent ingestion on postnatal day 15 in rats. Physiol. Behav. 62, 871-874]. It is possible that the equivalent inhibition of intake by the oil preloads on P12 resulted from the failure of the preabsorptive sensory properties of the preloads to be discriminated by peripheral or central sensory mechanisms. To investigate this possibility, we administered equivolumetric gastric preloads of 25% corn oil and 25% mineral oil to pups on P9-12 and counted the number of c-Fos-like immunoreactive (CFLI) cells in central sites that are activated by food intake and postingestive preabsortive mechanisms in adult rats and in pups on P10-11. The major result was that preloads of 25% corn oil and 25% mineral oil that produced equivalent inhibition of II intake produced differential increases of CFLI cells in the forebrain and hindbrain. Specifically, preloads of corn oil increased the number of CFLI cells in the caudal Nucleus Tractus Solitarius significantly more than preloads of mineral oil. Furthermore, preloads of corn oil increased the number of CFLI cells in the Paraventricular and Supraoptic nuclei, but preloads of mineral oil did not. This differential pattern of increases of CFLI cells is evidence that the brain discriminates the preabsorptive sensory properties of preloads of corn oil and mineral oil on P9-12.

Two different putative genetic animal models of childhood depression.

BACKGROUND: In an attempt to model childhood depression, we examined whether existing genetic animal models of depression in adult rats are also valid in prepubertal rats. METHODS: Two different "depressed" rat lines were studied: the Flinders Sensitive Line (FSL) and their controls, Sprague-Dawley (SD); and the Wistar Kyoto (WKY) line and their controls, Wistar. We hypothesized that male prepubertal FSL and WKY rats would show increased swim test immobility and different patterns of social play and of basal plasma levels of corticosterone and adrenocorticotropic hormone (ACTH) compared with control rats. RESULTS: Prepubertal FSL and WKY rats exhibited significantly longer duration of immobility than control rats in the swim test. The FSL rats demonstrated significantly higher levels of social play behaviors and lower levels of corticosterone and ACTH compared with SD control rats, whereas WKY rats demonstrated significantly lower levels of social play behaviors and higher plasma levels of corticosterone and ACTH compared with Wistar control rats. CONCLUSIONS: The results might suggest that prepubertal FSL and WKY rats are both putative genetic animal models of childhood depression, exhibiting separate patterns and symptoms of childhood depression.

Mother-Infant Interactions in Rats Lacking CCK(A) Receptors.

Mediation of mother-infant interactions by the brain-gut peptide cholecystokinin (CCK) was examined by observing behavior of Otsuka Long-Evans Tokushima Fatty (OLETF) rats, which lack functional CCKA receptors because of a genetic abnormality. OLETF (n = 10) and control (Long-Evans Tokushima Otsuka [LETO] n = 10) dams interacted with 1 pup of each line on Postpartum Days 6-9. OLETF pups received more body and anogenital licking and emitted substantially more ultrasonic vocalizations than LETO pups. OLETF dams carried pups less frequently and showed a nursing position more frequently than LETO dams. No significant Pup X Dam Line interactions or line differences in dams' activity were detected. The results provide convergent validity to previous pharmacological studies implicating CCK mediation of both infant and maternal behavior.

Traumatic brain injury - modeling neuropsychiatric symptoms in rodents.

Each year in the US, ∼1.5 million people sustain a traumatic brain injury (TBI). Victims of TBI can suffer from chronic post-TBI symptoms, such as sensory and motor deficits, cognitive impairments including problems with memory, learning, and attention, and neuropsychiatric symptoms such as depression, anxiety, irritability, aggression, and suicidal rumination. Although partially associated with the site and severity of injury, the biological mechanisms associated with many of these symptoms - and why some patients experience differing assortments of persistent maladies - are largely unknown. The use of animal models is a promising strategy for elucidation of the mechanisms of impairment and treatment, and learning, memory, sensory, and motor tests have widespread utility in rodent models of TBI and psychopharmacology. Comparatively, behavioral tests for the evaluation of neuropsychiatric symptomatology are rarely employed in animal models of TBI and, as determined in this review, the results have been inconsistent. Animal behavioral studies contribute to the understanding of the biological mechanisms by which TBI is associated with neurobehavioral symptoms and offer a powerful means for pre-clinical treatment validation. Therefore, further exploration of the utility of animal behavioral tests for the study of injury mechanisms and therapeutic strategies for the alleviation of emotional symptoms are relevant and essential.

The female urine sniffing test: a novel approach for assessing reward-seeking behavior in rodents.

BACKGROUND: Abnormal hedonic behavior is a key feature of many psychiatric disorders. Several paradigms measure reward-seeking behavior in rodents, but each has limitations. We describe a novel approach for monitoring reward-seeking behavior in rodents: sniffing of estrus female urine by male mice, along with number of ultrasonic vocalizations (USVs) emitted during the test. METHODS: The female urine sniffing test (FUST) was designed to monitor reward-seeking activity in rodents together with tests of helplessness and sweet solution preference. USVs and dopamine release from the nucleus accumbens (NAc) were recorded. Sniffing activity was measured in 1) manipulation-naive C57BL/6J and 129S1/SVImJ mice and Wistar-Kyoto rats; 2) stressed mice; 3) two groups of mice that underwent the learned helplessness paradigm-one untreated, and one treated with the SSRI citalopram; and 4) GluR6 knockout mice, known to display lithium-responsive, mania-related behaviors. RESULTS: Males from all three strains spent significantly longer sniffing female urine than sniffing water. Males emitted USVs and showed significantly elevated NAc dopamine levels while sniffing urine. Foot-shock stress significantly reduced female urine sniffing time. Compared with mice that did not undergo the LH paradigm, LH males spent less time sniffing female urine, and citalopram treatment alleviated this reduction. Compared with their wildtype littermates, GluR6KO males sniffed female urine longer and showed enhanced saccharin preference. CONCLUSIONS: In rodents, sniffing female urine is a preferred activity accompanied by biological changes previously linked to reward-seeking activities. The FUST is sensitive to behavioral and genetic manipulation and to relevant drug treatment.

Reverse translational strategies for developing animal models of bipolar disorder.

Bipolar disorder (BD) affects a significant portion of the population of the world, yet there has been limited success in developing novel treatments for the disorder. One of the major reasons for this dearth is the absence of suitable animal models for BD. Traditionally, animal models of human phenomena have been evaluated based on similarity to the human syndrome, response to appropriately corresponding medications, and the degree to which a model supports a common mechanistic theory between the human disorder and the model itself. The following review emphasizes the use of 'reverse translation', drawing on patient-based findings to develop suitable animal models for BD. We highlight some examples of this strategy, emphasizing their construct validity as a starting point. These studies have produced informative models that have altered the expression of genes/pathways implicated in BD, including the point mutation D181A of mouse mitochondrial DNA polymerase (POLG), glutamate receptor 6 (GluR6), Clock, extracellular regulated kinase 1 (ERK1), glycogen synthase kinase-3beta (GSK-3beta), B-cell lymphoma 2 (Bcl-2) and Bcl-2-associated athanogene (BAG-1). These studies demonstrate that this method is useful, viable and deserves attention in new efforts to generate animal models of BD.