Automated analysis of behavior: A computer‐controlled system for drug screening and the investigation of learning

Efforts to understand cognition will be greatly facilitated by computerized systems that enable the automated analysis of animal behavior. A number of controversies in the invertebrate learning field have resulted from difficulties inherent in manual experiments. Driven by the necessity to overcome these problems during investigation of neural function in planarian flatworms and frog larvae, we designed and developed a prototype for an inexpensive, flexible system that enables automated control and analysis of behavior and learning. Applicable to a variety of small animals such as flatworms and zebrafish, this system allows automated analysis of innate behavior, as well as of learning and memory in a plethora of conditioning paradigms. We present here the schematics of a basic prototype, which overcomes experimenter effects and operator tedium, enabling a large number of animals to be analyzed with transparent on‐line access to primary data. A scaled‐up version of this technology represents an efficient methodology to screen pharmacological and genetic libraries for novel neuroactive reagents of basic and biomedical relevance. © 2006 Wiley Periodicals, Inc. J Neurobiol, 2006     

Behavioral signatures of structured feature detection during courtship in Drosophila

1. Download : Download high-res image (193KB)
2. Download : Download full-size image

     

Automated identification of social interaction criteria in Drosophila melanogaster

The study of social behaviour within groups has relied on fixed definitions of an ‘interaction’. Criteria used in these definitions often involve a subjectively defined cut-off value for proximity, orientation and time (e.g. courtship, aggression and social interaction networks) and the same numerical values for these criteria are applied to all of the treatment groups within an experiment. One universal definition of an interaction could misidentify interactions within groups that differ in life histories, study treatments and/or genetic mutations. Here, we present an automated method for determining the values of interaction criteria using a pre-defined rule set rather than pre-defined values. We use this approach and show changing social behaviours in different manipulations of Drosophila melanogaster. We also show that chemosensory cues are an important modality of social spacing and interaction. This method will allow a more robust analysis of the properties of interacting groups, while helping us understand how specific groups regulate their social interaction space.     

Neural circuit mechanisms linking courtship and reward in Drosophila males

1. Download : Download high-res image (234KB)
2. Download : Download full-size image

     

Mushroom body ablation impairs short-term memory and long-term memory of courtship conditioning in Drosophila melanogaster

We have evaluated the role of the Drosophila mushroom bodies (MBs) in courtship conditioning, in which experience with mated females causes males to reduce their courtship toward virgins (Siegel and Hall, 1979). Whereas previous studies indicated that MB ablation abolished learning in an olfactory conditioning paradigm (deBelle and Heisenberg, 1994), MB-ablated males were able to learn in the courtship paradigm. They resumed courting at naive levels within 30 min after training, however, while the courtship of control males remained depressed 1 hr after training. We also describe a novel courtship conditioning paradigm that established long-term memory, lasting 9 days. In MB-ablated males, memory dissipated completely within 1 day. Our results indicate that the MBs are not required for learning and immediate recall of courtship conditioning but are required for consolidation of short-term and long-term associative memories.     

Aversive phototaxic suppression: evaluation of a short‐term memory assay in Drosophila melanogaster

Drosophila melanogaster is increasingly being used to model human conditions that are associated with cognitive deficits including fragile‐X syndrome, Alzheimer’s disease, Parkinson’s disease, sleep loss, etc. With few exceptions, cognitive abilities that are known to be modified in these conditions in humans have not been evaluated in fly models. One reason is the absence of a simple, inexpensive and reliable behavioral assay that can be used by laboratories that are not expert in learning and memory. Aversive phototaxic suppression (APS) is a simple assay in which flies learn to avoid light that is paired with an aversive stimulus (quinine/humidity). However, questions remain about whether the change in the fly’s behavior reflects learning an association between light and quinine/humidity or whether the change in behavior is because of nonassociative effects of habituation and/or sensitization. We evaluated potential effects of sensitization and habituation on behavior in the T‐maze and conducted a series of yoked control experiments to further exclude nonassociative effects and determine whether this task evaluates operant learning. Together these experiments indicate that a fly must associate the light with quinine/humidity to successfully complete the task. Next, we show that five classic memory mutants are deficient in this assay. Finally, we evaluate performance in a fly model of neurodegenerative disorders associated with the accumulation of Tau. These data indicate that APS is a simple and effective assay that can be used to evaluate fly models of human conditions associated with cognitive deficits.     

No trade-off between learning ability and parasitoid resistance in Drosophila melanogaster

Learning ability and immunity to parasites are linked at the physiological level in several insect species. The aim of this work was to investigate the relationship between learning and immunity at an evolutionary level. We tested whether selection for improved learning ability in Drosophila melanogaster led to changes in parasitoid resistance as a correlated response. Similarly, we assayed whether selection for better parasitoid resistance led to a change in learning ability. There was no significant difference between selected and control lines in either case; the estimated confidence intervals for the differences indicate that a trade-off relationship is unlikely.     

昆虫求偶行为在分类学上的应用

昆虫求偶行为是两性间相互吸引并完成各种生殖活动的一种定向行为,其形式多样,存在种的特异性。不同物种其求偶行为不同,以此来帮助区分特定类群近缘种、识别生物学种显得非常有意义。本文综述了性信息素求偶行为、鸣声求偶行为、闪光求偶行为以及献礼求偶行为在分类学上的应用。     

Deletion of the Drosophila neuronal gene found in neurons disrupts brain anatomy and male courtship

The fne (found‐in‐neurons) locus encodes one of the three paralogs of the ELAV gene family of Drosophila melanogaster. Members of this family are found throughout metazoans and encode RNA‐binding proteins with primarily neuronal localization, but with remarkably diverse functions given their high level of amino acid sequence conservation. The first identified member of the family, elav of Drosophila is a vital gene. Mutations in the second Drosophila elav paralog, rbp9, are viable but female sterile. No alleles of fne were previously available. FNE protein is normally present in the cytoplasm of all neurons throughout development. Here we describe the generation and characterization of fne^null mutations by homologous recombination. In contrast to elav and similar to rbp9, fne^null mutants are viable, but exhibit a specific and fully penetrant fusion of the β‐lobes in their mushroom bodies (MB), a paired neuropil of the central brain involved in a variety of complex behaviors. Mutant males have reduced courtship indices, but normal short‐ and long‐term courtship memory. Our data show that fne has specific functions which are non‐overlapping with the other two family members, namely in courtship behavior and in the development of the adult MB. The data further show that courtship memory does not require intact β‐lobes in the MB.     

Reassessment of the effect of biological rhythm mutations on learning in Drosophila melanogaster

A link between learning deficits and circadian period-lengthening mutations in Drosophila melanogaster previously has been reported. Mutant long-period males performed poorly in two learning assays involving experience-dependent courtship inhibition. In one, normal males that have courted fertilized females subsequently show courtship inhibition with virgin females. In the other, normal males that have courted sexually immature males subsequently fail to court other immature males. Those results have been reassessed in an extended study of genetic variants involving the period gene. 1. Long-period perL1 males demonstrated poor conditioned courtship inhibition when exposed to fertilized females; they showed normal courtship conditioning when exposed to immature males. This could be due to a perL1-associated olfactory deficit with fertilized females, since perL1 males were unable to discriminate behaviorally between fertilized and virgin females. 2. Other long-period males, including perL2 males and transgenic perL1 males bearing a truncated form of the per+ gene, were conditioned normally by fertilized females. Thus, the courtship inhibition defect is specific to the perL1 mutant strain. 3. perL1 (and other per mutant) flies showed normal acquisition and retention of a classically conditioned olfactory avoidance response. 4. Results from a new conditioned courtship inhibition experiment are presented; males exposed to fertilized females during training showed further courtship inhibition during subsequent exposure to fertilized females. From the perspective of learning theory, this can be viewed as a savings experiment.     

The effect of learning on experimental evolution of resource preference in Drosophila melanogaster

Abstract Learning is thought to be adaptive in variable environments, whereas constant, predictable environments are supposed to favor unconditional, genetically fixed responses. A dichotomous view of behavior as either learned or innate ignores a potential evolutionary interaction between the learned and innate components of a behavioral response. We addressed this interaction in the context of oviposition substrate choice in Drosophila melanogaster, asking two main questions. First, will learning also evolve in a constant environment in which it always pays to show the same choice? Second, how does an opportunity to learn affect the evolution of the innate (genetic) component of oviposition substrate choice? We exposed experimental populations to four selection regimes, involving selection on oviposition substrate preference (an orange versus a pineapple medium). In two selection regimes the flies were selected for preference either for the orange medium, or for the pineapple medium. In the remaining two selection regimes the flies were also selected for preference for either orange or pineapple, but additionally could use past experience (aversion learning) to decide which medium it paid to avoid. Lines exposed to the latter selection regimes evolved improved learning ability, indicating that learning may be advantageous even if the same behavioral response is favored every generation. Furthermore, of the two selection regimes that favored oviposition on the pineapple medium, the regime that allowed for learning led to the evolution of a stronger innate preference for pineapple, than the regime that did not allow for learning. In contrast, of the two regimes that selected for oviposition on the orange medium, the one that allowed for learning led to a smaller evolutionary change of the innate preference. Thus, an opportunity to learn facilitated the evolution of innate preference under selection for preference for pineapple, but hindered it under selection for preference for orange. We discuss possible mechanisms for this effect.     

Relationship between visual learning/memory ability and brain cAMP level in Drosophila

Ｄｒｏｓｏｐｈｉｌａｈａｓｂｅｃｏｍｅａｎｉｍｐｏｒｔａｎｔｅｘｐｅｒｉｍｅｎｔａｌａｎｉｍａｌｍｏｄｅｌｂｅｃａｕｓｅｏｆｉｔｓｔｗｏａｄｖａｎｔａｇｅｓ,ｉ.ｅ.ｉｔｓｒｅｌａｔｉｖｅｌｙｃｌｅａｒｇｅｎｅｔｉｃｂａｃｋｇｒｏｕｎｄａｎｄａｖａｒｉｅｔｙｏｆａｓｓｏｃｉａｔｉｖｅｌｅａｒｎｉｎｇａｂｉｌｉｔｉｅｓ.Ｓｉｎｇｌｅｇｅｎｅｏｌｆａｃｔｏｒｙｌｅａｒｎｉｎｇｍｕｔａｎｔｓｈａｖｅｂｅｅｎｉｓｏｌａｔｅｄ.ＳｏｍｅｏｆｔｈｅｍａｒｅｒｅｌａｔｅｄｔｏｔｈｅｃＡＭＰｓｉｇｎａｌｔｒａｎｓｄ…     

Esterase-6 and the pheromonal effects of cis-vaccenyl acetate in Drosophila melanogaster

In Drosophila melanogaster the polymorphic enzyme Est-6 and a male specific lipid, cis-vaccenyl acetate (cVA), are components of the seminal fluid. It has been suggested that cVA could be a physiological substrate for Est-6, constituting, together with the hypothetical hydrolytic product, cis-vaccenyl alcohol (cVOH), a pheromonal system active in the regulation of the sexual attractiveness of mated females. However, some doubt exists concerning the physiological conversion of cVA to cVOH in females. Est-6 is also known to affect sperm motility, sperm use and female fecundity. The results of a study on qualitative and quantitative effects of topical treatment of females with cis-vaccenyl acetate and cis-vaccenyl alcohol on courtship behaviour are presented. The inhibition of courtship produced by cVOH was more persistent; this was possibly related to its lower volatility. The hypothesis was tested that different Est-6 allozymes could produce different amounts of cis-vaccenyl alcohol in vivo by differential hydrolysis of cis-vaccenyl acetate, and thus affect the timing of remating. For this purpose highly homogeneous Est-6^S and Est-6^F homozygous lines, with almost certainly identical levels of Est-6 and cVA, were used. They were obtained after 100 generations of inbreeding and selection of the heterozygotes at each generation. The Est-6^S and Est-6^F products of the Est-6 structural gene did not differentially affect the timing of remating. This could imply that the two allozymes have similar catalytic properties. However, such results could be obtained even if cVA is not converted to cVOH in vivo by Est-6. The effects of Est-6 allozymes on sperm use after remating were also investigated. The general effect of second males was a very high elimination of first male fertilizations. No evidence for a different contribution of the two allozymes to the fitness through sperm predominance was obtained. Remating and sperm predominance experiments were performed at three temperatures: 18, 25 and 30° C. A more general—not species specific—role of cis-vaccenyl esters is suggested by preliminary results on their presence in the ejaculate of other Drosophila species.     

牛磺酸对学习记忆影响的研究现状

牛磺酸作为一种条件必须氨基酸对学习记忆能力的影响受到越来越多的关注,许多研究证明适量牛磺酸的添加可明显提高大鼠的记忆能力。通过牛磺酸对神经细胞、基因、激素、离子、受体、酶类等的作用,对其影响学习记忆方面的研究现状作出综述。     

Genetically idiosyncratic responses of Drosophila melanogaster populations to selection for improved learning ability

To what extent is adaptive evolution over short timescales repeatable? To address this question, we studied the performance of crosses between replicate Drosophila melanogaster lines previously subject to selection for improved learning response in the context of oviposition substrate choice. Of the 10 pairwise F₁ crosses among the five selection lines, four performed in the original learning assay similarly to the parental lines, whereas the remaining six showed learning scores significantly below the average of the parental lines. In particular, four F₁ crosses (three involving the same line) showed no detectable learning, on a par with unselected control lines. This indicates that the response to selection in some lines involved allelic substitutions at different loci. Additional assays of crosses between two selection lines indicated that the loss of performance in hybrids generalized to another type of learning assay, and held for both short‐ and long‐term memory. Joint analysis of first‐ and second‐generation crosses between these two lines supported the hypothesis that the response to selection in these different lines was based on the spread of recessive alleles at different loci. These results show that the evolutionary trajectories of populations of the same origin subject to uniform selection may sometimes diverge over very short evolutionary timescales.     

学习记忆相关基因研究进展

学习记忆是脑的重要功能之一,与学习记忆相关的基因很多,它们通过影响突触功能、信号转导、转录和翻译控制、能量代谢等途径进行学习记忆行为的调控。研究相关基因可为阐明学习记忆的分子机制和遗传机制奠定基础。