Behavioral modification in choice process of Drosophila

Because of its clear genetic and developmental background, diversity of behavioral paradigms and neuroanatomy of the brain, Drosophila has become an important animal model for studying genetic, molecular and cellular bases of learning and memory[1]. Extensive research has explored the visual operant conditioning of Drosophila and related molecular bases[2—8]; recently, researchers began to address cognition-like functions and involved neural substrates[9—11]. In these studies, behavioral ana…     

Behavioral modification in choice process of<Emphasis Type="Italic">Drosophila</Emphasis>

In visual operant conditioning ofDrosophila at the flight simulator, only motor output of flies—yaw torque—is recorded, which is involved in the conditioning process. The current study used a newly-designed data analysis method to study the torque distribution ofDrosophila. Modification of torque distribution represents the effects of operant conditioning on flies’ behavioral mode. Earlier works^[10] showed that, when facing contradictory visual cues, flies could make choices based upon the relative weightiness of different cues, and it was demonstrated that mushroom bodies might play an important role in such choice behavior. The new “torque-position map” method was used to explore the CS-US associative learning and choice behavior inDrosophila from the aspect of its behavioral mode. Finally, this work also discussed various possible neural bases involved in visual associative learning, choice processing and modification processing of the behavioral mode in the visual operant conditioning ofDrosophila.     

Associative learning and memory in Drosophila: beyond olfactory conditioning

The associative learning abilities of the fruit fly, Drosophila melanogaster, have been demonstrated in both classical and operant conditioning paradigms. Efforts to identify the neural pathways and cellular mechanisms of learning have focused largely on olfactory classical conditioning. Results derived from various genetic and molecular manipulations provide considerable evidence that this form of associative learning depends critically on neural activity and cAMP signaling in brain neuropil structures called mushroom bodies. Three other behavioral learning paradigms in Drosophila serve as the main subject of this review. These are (1) visual and motor learning of flies tethered in a flight simulator, (2) a form of spatial learning that is independent of visual and olfactory cues, and (3) experience-dependent changes in male courtship behavior. The present evidence suggests that at least some of these modes of learning are independent of mushroom bodies. Applying targeted genetic manipulations to these behavioral paradigms should allow for a more comprehensive understanding of neural mechanisms responsible for diverse forms of associative learning and memory.     

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

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苯甲醛对果蝇视觉联想记忆的阻断

采用飞行模拟系统,以视觉模式为线索、热惩罚为负强化因子,对于在不同发育时期经受苯甲醛处理过的果蝇的视觉飞行定向条件化进行了检验。苯甲醛气味分别作用于果蝇幼虫和成虫阶段,将阻断果蝇成虫建立视觉联想记忆的能力;雌性果蝇在处女期对苯甲醛气味的接触,会阻断其子代建立视觉联想记忆,这种视觉联想记忆的能力可以通过对其子代连续３代的正常饲养而逐渐得到恢复。     

Operant conditioning in Drosophila melanogaster wild-type and learning mutants with defects in the cyclic AMP metabolism

Wild-type Drosophila melanogaster and the learning mutants dunce, amnesiac and rutabaga, were tested using a new operant conditioning paradigm for single flies. All strains are able to learn to different extents, but no evidence of memory was found in the mutants amnesiac and rutabaga, while dunce has a reduced but extended memory. The relationship between this characteristic and cAMP levels are discussed. The three mutants have previously been shown, using classical conditioning paradigms to be deficient in olfactory learning and/or memory, and show reduced visual learning. The variability of the response of the mutants in the different paradigms is discussed in relation to the generality of the Aplysia model of the cellular mechanism underlying learning. In the operant conditioning paradigm described here, 93% of the wild-type flies learned to criterion. The performance of individual flies was consistent.     

Relationship between visual learning/memory ability and brain cAMP level inDrosophila

Using the flight simulator system, the operant conditioned visual flight orientation behavior inDrosophila was studied. It was demonstrated that the visual learning performance is associated with age; flies learn more reliably at 3–4 days than at 1–2 days of age; the cAMP level of brain is also increasing with age; the brain cAMP content of nonlearner flies of wild type is much higher than that of normal flies; the cAMP level of brain increased abnormally after being fed with caffeine, and the learning performance decred. These results imply that a moderate range of cAMP level is necessary for the visual learning and memory pmess. Abnody high or low level of cAMP causes defects of leaming and memory ability.     

Classical reward conditioning in Drosophila melanogaster

Negatively reinforced olfactory conditioning has been widely employed to identify learning and memory genes, signal transduction pathways and neural circuitry in Drosophila. To delineate the molecular and cellular processes underlying reward-mediated learning and memory, we developed a novel assay system for positively reinforced olfactory conditioning. In this assay, flies were involuntarily exposed to the appetitive unconditioned stimulus sucrose along with a conditioned stimulus odour during training and their preference for the odour previously associated with sucrose was measured to assess learning and memory capacities. After one training session, wild-type Canton S flies displayed reliable performance, which was enhanced after two training cycles with 1-min or 15-min inter-training intervals. Higher performance scores were also obtained with increasing sucrose concentration. Memory in Canton S flies decayed slowly when measured at 30 min, 1 h and 3 h after training; whereas, it had declined significantly at 6 h and 12 h post-training. When learning mutant t beta h flies, which are deficient in octopamine, were challenged, they exhibited poor performance, validating the utility of this assay. As the Drosophila model offers vast genetic and transgenic resources, the new appetitive conditioning described here provides a useful tool with which to elucidate the molecular and cellular underpinnings of reward learning and memory. Similar to negatively reinforced conditioning, this reward conditioning represents classical olfactory conditioning. Thus, comparative analyses of learning and memory mutants in two assays may help identify the molecular and cellular components that are specific to the unconditioned stimulus information used in conditioning.     

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

Using the flight simulator system, the operant conditioned visual flight orientation behavior inDrosophila was studied. It was demonstrated that the visual learning performance is associated with age; flies learn more reliably at 3–4 days than at 1–2 days of age; the cAMP level of brain is also increasing with age; the brain cAMP content of nonlearner flies of wild type is much higher than that of normal flies; the cAMP level of brain increased abnormally after being fed with caffeine, and the learning performance decred. These results imply that a moderate range of cAMP level is necessary for the visual learning and memory pmess. Abnody high or low level of cAMP causes defects of leaming and memory ability. Project supported by the National Natural Science Foundation of China rant No. 69435013) and the National Fundarnental Research Programme in China     

Attention-like processes underlying optomotor performance in a Drosophila choice maze

The authors present a novel paradigm for studying visual responses in Drosophila. An eight-level choice maze was found to reliably segregate fly populations according to their responses to moving stripes displayed on a computer screen. Visual responsiveness was robust in wild-type flies, and performance depended on salience effects such as stimulus color and speed. Analysis of individual fly choices in the maze revealed that stereotypy, or choice persistence, contributed significantly to a strain's performance. On the basis of these observations, the authors bred wild-type flies for divergent visual phenotypes by selecting individual flies displaying extreme stereotypy. Selected flies alternated less often in the sequential choice maze than unselected flies, showing that stereotypy could evolve across generations. The authors found that selection for increased stereotypy impaired flies' responsiveness to competing stimuli in tests for attention-like behavior in the maze. Visual selective attention was further investigated by electrophysiology, and it was found that increased stereotypy also impaired responsiveness to competing stimuli at the level of brain activity. Combined results present a comprehensive approach to studying visual responses in Drosophila, and show that behavioral performance involves attention-like processes that are variable among individuals and thus sensitive to artificial selection.     

Olfactory conditioning of proboscis activity in Drosophila melanogaster

Olfactory learning and memory processes in Drosophila have been well investigated with aversive conditioning, but appetitive conditioning has rarely been documented. Here, we report for the first time individual olfactory conditioning of proboscis activity in restrained Drosophila melanogaster. The protocol was adapted from those developed for proboscis extension conditioning in the honeybee Apis mellifera. After establishing a scale of small proboscis movements necessary to characterize responses to olfactory stimulation, we applied Pavlovian conditioning, with five trials consisting of paired presentation of a banana odour and a sucrose reward. Drosophila showed conditioned proboscis activity to the odour, with a twofold increase of percentage of responses after the first trial. No change occurred in flies experiencing unpaired presentations of the stimuli, confirming an associative basis for this form of olfactory learning. The adenylyl cyclase mutant rutabaga did not exhibit learning in this paradigm. This protocol generated at least a short-term memory of 15 min, but no significant associative memory was detected at 1 h. We also showed that learning performance was dependent on food motivation, by comparing flies subjected to different starvation regimes.     

Deficient protein kinase C activity in turnip, a Drosophila learning mutant

The Drosophila mutant turnip was initially isolated based on poor learning performance (Quinn, W.G., Sziber, P.P., and Booker, R. (1979) Nature 277, 212-214). Here we show that turnip is dramatically reduced in protein kinase C (PKC) activity. In addition, turnip flies are deficient in phosphorylation of a 76-kDa head membrane protein (hereafter pp76) which is a major substrate for protein kinase C in homogenates of wild-type flies. Reduced PKC activity, defective pp76 phosphorylation, and most of turnip's learning deficiency co-map genetically to a region on the X-chromosome, 18A5-18D1-2, spanned by the deletion Df(1)JA27. Apparently turnip+ is not a structural gene for PKC because Drosophila PKC genes map elsewhere in the genome. Our results suggest that turnip gene product is required for activation of PKC and that PKC plays a role in associative learning in Drosophila.     

Basic organization of operant behavior as revealed in Drosophila flight orientation

Summary Operant behavior is studied in tethered Drosophila flies using visual motion, heat or odour as operandum and yaw torque, thrust or direction of flight as operans in various combinations (Fig. 1). On the basis of these results a conceptual framework of operant behavior is proposed: (1) It requires a goal (desired state) of which the actual state deviates. (2) To attain the goal a range of motor programs is activated (initiating activity, see Fig. 7). (3) Efference copies of the motor programs are compared to the sensory input referring to the deviation from the desired state (e.g. by cross-correlation). (4) In case of a significant coincidence the respective motor program is used to modify the sensory input in the direction towards the goal. (5) Consistent control of a sensory stimulus by a behavior may lead to a more permanent behavioral change (conditioning). In this scheme operant activity (1–4) and operant conditioning (1–5) are distinguished.Abbreviations ALU arbitrary length unit - d horizontal angular width of visual pattern - IR infrared - SEM standard error of the means - T yaw torque - Th thrust - performance index - horizontal angle between visual pattern position and longitudinal body axis of the fly - vertical angular extension of visual pattern     

Intrinsic activity in the fly brain gates visual information during behavioral choices

The small insect brain is often described as an input/output system that executes reflex-like behaviors. It can also initiate neural activity and behaviors intrinsically, seen as spontaneous behaviors, different arousal states and sleep. However, less is known about how intrinsic activity in neural circuits affects sensory information processing in the insect brain and variability in behavior. Here, by simultaneously monitoring Drosophila's behavioral choices and brain activity in a flight simulator system, we identify intrinsic activity that is associated with the act of selecting between visual stimuli. We recorded neural output (multiunit action potentials and local field potentials) in the left and right optic lobes of a tethered flying Drosophila, while its attempts to follow visual motion (yaw torque) were measured by a torque meter. We show that when facing competing motion stimuli on its left and right, Drosophila typically generate large torque responses that flip from side to side. The delayed onset (0.1-1 s) and spontaneous switch-like dynamics of these responses, and the fact that the flies sometimes oppose the stimuli by flying straight, make this behavior different from the classic steering reflexes. Drosophila, thus, seem to choose one stimulus at a time and attempt to rotate toward its direction. With this behavior, the neural output of the optic lobes alternates; being augmented on the side chosen for body rotation and suppressed on the opposite side, even though the visual input to the fly eyes stays the same. Thus, the flow of information from the fly eyes is gated intrinsically. Such modulation can be noise-induced or intentional; with one possibility being that the fly brain highlights chosen information while ignoring the irrelevant, similar to what we know to occur in higher animals.     

The effect of removing the N-terminal extension of the Drosophila myosin regulatory light chain upon flight ability and the contractile dynamics of indirect flight muscle

The Drosophila myosin regulatory light chain (DMLC2) is homologous to MLC2s of vertebrate organisms, except for the presence of a unique 46-amino acid N-terminal extension. To study the role of the DMLC2 N-terminal extension in Drosophila flight muscle, we constructed a truncated form of the Dmlc2 gene lacking amino acids 2-46 (Dmlc2(Delta2-46)). The mutant gene was expressed in vivo, with no wild-type Dmlc2 gene expression, via P-element-mediated germline transformation. Expression of the truncated DMLC2 rescues the recessive lethality and dominant flightless phenotype of the Dmlc2 null, with no discernible effect on indirect flight muscle (IFM) sarcomere assembly. Homozygous Dmlc2(Delta2-46) flies have reduced IFM dynamic stiffness and elastic modulus at the frequency of maximum power output. The viscous modulus, a measure of the fly's ability to perform oscillatory work, was not significantly affected in Dmlc2(Delta2-46) IFM. In vivo flight performance measurements of Dmlc2(Delta2-46) flies using a visual closed-loop flight arena show deficits in maximum metabolic power (P(*)(CO(2))), mechanical power (P(*)(mech)), and flight force. However, mutant flies were capable of generating flight force levels comparable to body weight, thus enabling them to fly, albeit with diminished performance. The reduction in elastic modulus in Dmlc2(Delta2-46) skinned fibers is consistent with the N-terminal extension being a link between the thick and thin filaments that is parallel to the cross-bridges. Removal of this parallel link causes an unfavorable shift in the resonant properties of the flight system, thus leading to attenuated flight performance.     

cAMP-dependent protein kinase and the disruption of learning in transgenic flies.

A molecular genetic approach was used to test for a role of cAMP-dependent protein kinase (PKA) in learning and memory in Drosophila. We used genes encoding a peptide inhibitor of PKA, an N-terminal regulatory subunit fragment containing a pseudosubstrate inhibitory domain, and a wild-type catalytic subunit. These dominantly acting genes were placed under control of the hsp70 promoter and transformed into wild-type flies. Induction of the transgenes by 1 hr heat shock results in overproduction of their RNA in adult flies. The same heat shock treatment disrupts the ability of the flies to learn in an odor discrimination task reinforced with electric shock. The results demonstrate the involvement of PKA in the associative learning of Drosophila.     

What can we teach Lymnaea and what can Lymnaea teach us?

This review describes the advantages of adopting a molluscan complementary model, the freshwater snail Lymnaea stagnalis, to study the neural basis of learning and memory in appetitive and avoidance classical conditioning; as well as operant conditioning of its aerial respiratory and escape behaviour. We firstly explored ‘what we can teach Lymnaea’ by discussing a variety of sensitive, solid, easily reproducible and simple behavioural tests that have been used to uncover the memory abilities of this model system. Answering this question will allow us to open new frontiers in neuroscience and behavioural research to enhance our understanding of how the nervous system mediates learning and memory. In fact, from a translational perspective, Lymnaea and its nervous system can help to understand the neural transformation pathways from behavioural output to sensory coding in more complex systems like the mammalian brain. Moving on to the second question: ‘what can Lymnaea teach us?’, it is now known that Lymnaea shares important associative learning characteristics with vertebrates, including stimulus generalization, generalization of extinction and discriminative learning, opening the possibility to use snails as animal models for neuroscience translational research.     

Golgi and genetic mosaic analyses of visual system mutants in Drosophila melanogaster

We have used a Golgi staining procedure in Drosophila melanogaster to examine the structure of individual neurons in the visual systems of the Canton-S wild-type strain, of flies expressing mutations at the Glued, rough, glass, and uneven loci, all of which affect the organization of the visual system, and of genetic mosaics involving the Glued and uneven loci. We have found that the structure of the neurons studied in the wild type is quite similar to that reported for other diptera and that the mutants studied evidence a variety of abnormalities in neuronal morphology, each mutant being characterized by a different spectrum of aberrations. The genetic mosaic analysis of the Glued and uneven loci showed that the structure of individual neurons in the optic lobes is profoundly influenced by the genotype of the cells projecting to that region from the compound eye but that the final form attained by a neuron is not solely controlled by that factor.     

The circadian basis of ovarian diapause regulation in Drosophila melanogaster: is the period gene causally involved in photoperiodic time measurement?

Females of a wild-type strain of Drosophila melanogaster (Canton-S), and of several clock mutants (period), were able to discriminate between diapause-inducing short days and diapause-averting long days with a well-defined critical daylength. The critical daylengths of a short-period mutant (pers) and a long-period mutant (perL2) were almost identical, both to each other and to that of Canton-S. The critical daylength of an arrhythmic mutant (perol), however, was about 3 hr shorter than that of Canton-S, and that of per- was about 5 hr shorter. Exposure of Canton-S females to Nanda-Hamner experiments, consisting of a 10-hr photophase coupled to a dark phase varying between 4 and 74 hr, showed (1) that the photoperiodic clock in D. melanogaster measures nightlength rather than daylength, and (2) that photoperiodic time measurement is somehow based on (or affected by) constituent oscillators in the circadian system. Nanda-Hamner results for the period mutants all showed similar profiles regardless of genotype, or the presence or absence of per locus DNA. These results suggest that photoperiodic induction and locomotor activity do not share a common pacemaker in D. melanogaster, and that the per gene is not causally involved in nightlength measurement by the photoperiodic clock, although flies in which the per locus is missing (per-) or defective (perol) show an altered critical value.     

Distinct Acute Zones for Visual Stimuli in Different Visual Tasks in Drosophila

The fruit fly Drosophila melanogaster has a sophisticated visual system and exhibits complex visual behaviors. Visual responses, vision processing and higher cognitive processes in Drosophila have been studied extensively. However, little is known about whether the retinal location of visual stimuli can affect fruit fly performance in various visual tasks. We tested the response of wild-type Berlin flies to visual stimuli at several vertical locations. Three paradigms were used in our study: visual operant conditioning, visual object fixation and optomotor response. We observed an acute zone for visual feature memorization in the upper visual field when visual patterns were presented with a black background. However, when a white background was used, the acute zone was in the lower visual field. Similar to visual feature memorization, the best locations for visual object fixation and optomotor response to a single moving stripe were in the lower visual field with a white background and the upper visual field with a black background. The preferred location for the optomotor response to moving gratings was around the equator of the visual field. Our results suggest that different visual processing pathways are involved in different visual tasks and that there is a certain degree of overlap between the pathways for visual feature memorization, visual object fixation and optomotor response.