Learning and memory deficits upon TAU accumulation in Drosophila mushroom body neurons

Mutations in the neuronal-specific microtubule-binding protein TAU are associated with several dementias and neurodegenerative diseases. However, the effects of elevated TAU accumulation on behavioral plasticity are unknown. We report that directed expression of wild-type vertebrate and Drosophila TAU in adult mushroom body neurons, centers for olfactory learning and memory in Drosophila, strongly compromised associative olfactory learning and memory, but olfactory conditioning-relevant osmotactic and mechanosensory responses remained intact. In addition, TAU accumulation in mushroom body neurons did not result in detectable neurodegeneration or premature death. Therefore, TAU-mediated structural or functional perturbation of the microtubular cytoskeleton in mushroom body neurons is likely causal of the behavioral deficit. These results indicate that behavioral plasticity decrements may be the earliest detectable manifestations of tauopathies.     

Roles for Drosophila mushroom body neurons in olfactory learning and memory

Olfactory learning assays in Drosophila have revealed that distinct brain structures known as mushroom bodies (MBs) are critical for the associative learning and memory of olfactory stimuli. However, the precise roles of the different neurons comprising the MBs are still under debate. The confusion surrounding the roles of the different neurons may be due, in part, to the use of different odors as conditioned stimuli in previous studies. We investigated the requirements for the different MB neurons, specifically the alpha/beta versus the gamma neurons, and whether olfactory learning is supported by different subsets of MB neurons irrespective of the odors used as conditioned stimuli. We expressed the rutabaga (rut)-encoded adenylyl cyclase in either the gamma or alpha/beta neurons and examined the effects on restoring olfactory associative learning and memory of rut mutant flies. We also expressed a temperature-sensitive shibire (shi) transgene in these neuron sets and examined the effects of disrupting synaptic vesicle recycling on Drosophila olfactory learning. Our results indicate that although we did not detect odor-pair-specific learning using GAL4 drivers that primarily express in gamma neurons, expression of the transgenes in a subset of alpha/beta neurons resulted in both odor-pair-specific rescue of the rut defect as well as odor-pair-specific disruption of learning using shi(ts1).     

Mushroom Bodies Suppress Locomotor Activity in Drosophila melanogaster

Locomotor activity of single, freely walking flies in small tubes is analyzed in the time domain of several hours. To assess the influence of the mushroom bodies on walking activity, three independent noninvasive methods interfering with mushroom body function are applied: chemical ablation of the mushroom body precursor cells; a mutant affecting Kenyon cell differentiation (mushroom body miniature¹); and the targeted expression of the catalytic subunit of tetanus toxin in subsets of Kenyon cells. All groups of flies with mushroom body defects show an elevated level of total walking activity. This increase is attributable to the slower and less complete attenuation of activity during the experiment. Walking activity in normal and mushroom body-deficient flies is clustered in active phases (bouts) and rest periods (pauses). Neither the initiation nor the internal structure, but solely the termination of bouts seems to be affected by the mushroom body defects. How this finding relates to the well-documented role of the mushroom bodies in olfactory learning and memory remains to be understood.     

Mushroom bodies of the honeybee brain show cell population-specific plasticity in expression of amine-receptor genes

Dopamine and octopamine released in the mushroom bodies of the insect brain play a critical role in the formation of aversive and appetitive memories, respectively. As recent evidence suggests a complex relationship between the effects of these two amines on the output of mushroom body circuits, we compared the expression of dopamine- and octopamine-receptor genes in three major subpopulations of mushroom body intrinsic neurons (Kenyon cells). Using the brain of the honeybee, Apis mellifera, we found that expression of amine-receptor genes differs markedly across Kenyon cell subpopulations. We found, in addition, that levels of expression of these genes change dramatically during the lifetime of the bee and that shifts in expression are cell population-specific. Differential expression of amine-receptor genes in mushroom body neurons and the plasticity that exists at this level are features largely ignored in current models of mushroom body function. However, our results are consistent with the growing body of evidence that short- and long-term olfactory memories form in different regions of the mushroom bodies of the brain and that there is functional compartmentalization of the modulatory inputs to this multifunctional brain center.     

Drosophila Mushroom Bodies Are Dispensable for Visual, Tactile, and Motor Learning

A total of 18 associative learning/memory tests have been applied to Drosophila melanogaster flies lacking mushroom bodies. Only in paradigms involving chemosensory cues as conditioned stimuli have flies been found to be compromised by a block in the mushroom body pathway. Among the learning tasks not requiring these structures are a case of motor learning (yaw torque/heat), a test of the fly’s spatial orientation in total darkness, conditioned courtship suppression by mated females, and nine different examples of visual learning. The latter used the reinforcers of heat, visual oscillations, mechanical shaking, or sucrose, and as conditioned stimuli, color, intensity contrast, as well as stationary and moving visual patterns. No forms of consolidated memory have been tested in mushroom body-less flies. With respect to short-term memory the mushroom bodies of Drosophila are specially required for chemosensory learning tasks, but not for associative learning and memory in general.     

Visual pattern memory requires foraging function in the central complex of Drosophila

The role of the foraging (for) gene, which encodes a cyclic guanosine-3',5'-monophosphate (cGMP)-dependent protein kinase (PKG), in food-search behavior in Drosophila has been intensively studied. However, its functions in other complex behaviors have not been well-characterized. Here, we show experimentally in Drosophila that the for gene is required in the operant visual learning paradigm. Visual pattern memory was normal in a natural variant rover (for(R)) but was impaired in another natural variant sitter (for(S)), which has a lower PKG level. Memory defects in for(S) flies could be rescued by either constitutive or adult-limited expression of for in the fan-shaped body. Interestingly, we showed that such rescue also occurred when for was expressed in the ellipsoid body. Additionally, expression of for in the fifth layer of the fan-shaped body restored sufficient memory for the pattern parameter "elevation" but not for "contour orientation," whereas expression of for in the ellipsoid body restored sufficient memory for both parameters. Our study defines a Drosophila model for further understanding the role of cGMP-PKG signaling in associative learning/memory and the neural circuit underlying this for-dependent visual pattern memory.     

Metamorphosis of the Mushroom Bodies; Large-Scale Rearrangements of the Neural Substrates for Associative Learning and Memory in Drosophila

Paired brain centers known as mushroom bodies are key features of the circuitry for insect associative learning, especially when evoked by olfactory cues. Mushroom bodies have an embryonic origin, and unlike most other brain structures they exhibit developmental continuity, being prominent components of both the larval and the adult CNS. Here, we use cell-type-specific markers, provided by the P{GAL4} enhancer trap system, to follow specific subsets of mushroom body intrinsic and extrinsic neurons from the larval to the adult stage. We find marked structural differences between the larval and adult mushroom bodies, arising as the consequence of large-scale reorganization during metamorphosis. Extensive, though incomplete, degradation of the larval structure is followed by establishment of adult specific α and β lobes. Kenyon cells of embryonic origin, by contrast, were found to project selectively to the adult γ lobe. We propose that the γ lobe stores information of relevance to both developmental stages, whereas the α and β lobes have uniquely adult roles.     

Tripartite Mushroom Body Architecture Revealed by Antigenic Markers

We have explored the organization of the axonal lobes in Drosophila mushroom bodies by using a panel of immunohistochemical markers. These markers consist of antibodies to eight proteins expressed preferentially in the mushroom bodies: DAMB, DCO, DRK, FASII, LEO, OAMB, PKA RII, and RUT. Previous to this work, four axonal lobes, two projecting dorsally (α and α′) and two medially (β and γ), had been described in Drosophila mushroom bodies. However, our analysis of immunohistochemically stained frontal and sagittal sections of the brain revealed three medially projecting lobes. The newly distinguished lobe, which we term β′, lies along the dorsal surface of β, just posterior to γ. In addition to resolving a fifth lobe, our studies revealed that there are specific lobe sets defined by equivalent marker expression levels. These sets are (1) the α and β lobes, (2) the α′ and β′ lobes, and (3) the γ lobe and heel (a lateral projection formed by a hairpin turn of some of the peduncle fibers). All of the markers we have examined are consistent with these three sets. Previous Golgi studies demonstrate that each mushroom body cell projects one axon that branches into a dorsal lobe and a medial lobe, or one unbranched axon that projects medially. Taken together with the lobe sets listed above, we propose that there are three major projection configurations of mushroom body cell axons: (1) one branch in the α and one in the β lobe, (2) one branch in the α′ and one in the β′ lobe, and (3) one unbranched axon projecting to the heel and the γ lobe. The fact that these neuron types exhibit differential expression levels of a number of mushroom body genes suggests that they may have corresponding functional differences. These functions may be conserved in the larvae, as several of these genes were expressed in larval and embryonic mushroom bodies as well. The basic mushroom body structure, including the denritic calyx, peduncle, and lobes, was already visible by the late stages of embryogenesis. With new insights into mushroom body organization, and the characterization of markers for developing mushroom bodies, we are beginning to understand how these structures form and function.     

Evolution, Discovery, and Interpretations of Arthropod Mushroom Bodies

Mushroom bodies are prominent neuropils found in annelids and in all arthropod groups except crustaceans. First explicitly identified in 1850, the mushroom bodies differ in size and complexity between taxa, as well as between different castes of a single species of social insect. These differences led some early biologists to suggest that the mushroom bodies endow an arthropod with intelligence or the ability to execute voluntary actions, as opposed to innate behaviors. Recent physiological studies and mutant analyses have led to divergent interpretations. One interpretation is that the mushroom bodies conditionally relay to higher protocerebral centers information about sensory stimuli and the context in which they occur. Another interpretation is that they play a central role in learning and memory. Anatomical studies suggest that arthropod mushroom bodies are predominately associated with olfactory pathways except in phylogenetically basal insects. The prominent olfactory input to the mushroom body calyces in more recent insect orders is an acquired character. An overview of the history of research on the mushroom bodies, as well as comparative and evolutionary considerations, provides a conceptual framework for discussing the roles of these neuropils.     

G(o) activation is required for both appetitive and aversive memory acquisition in Drosophila

Heterotrimeric G(o) is an abundant brain protein required for negatively reinforced short-term associative olfactory memory in Drosophila. G(o) is the only known substrate of the S1 subunit of pertussis toxin (PTX) in fly, and acute expression of PTX within the mushroom body neurons (MB) induces a reversible deficit in associative olfactory memory. We demonstrate here that the induction of PTX within the α/β and γ lobe MB neurons leads to impaired memory acquisition without affecting memory stability. The induction of PTX within these MB neurons also leads to a significant defect in an optimized positively reinforced short-term memory paradigm; however, this PTX-induced learning deficit is noticeably less severe than found with the negatively reinforced paradigm. Both negatively and positively reinforced memory phenotypes are rescued by the constitutive expression of G(o)α transgenes bearing the Cys(351)Ile mutation. Since this mutation renders the G(o) molecule insensitive to PTX, the results isolate the effect of PTX on both forms of olfactory associative learning to the inhibition of the G(o) activation.     

Protection from premature habituation requires functional mushroom bodies in Drosophila

Diminished responses to stimuli defined as habituation can serve as a gating mechanism for repetitive environmental cues with little predictive value and importance. We demonstrate that wild-type animals diminish their responses to electric shock stimuli with properties characteristic of short- and long-term habituation. We used spatially restricted abrogation of neurotransmission to identify brain areas involved in this behavioral response. We find that the mushroom bodies and, in particular, the α/β lobes appear to guard against habituating prematurely to repetitive electric shock stimuli. In addition to protection from premature habituation, the mushroom bodies are essential for spontaneous recovery and dishabituation. These results reveal a novel modulatory role of the mushroom bodies on responses to repetitive stimuli in agreement with and complementary to their established roles in olfactory learning and memory.     

正立和倒立的威胁性身体表情的识别

为探索身体表情的加工时程以及加工方式是否为构形加工,采用ERP技术考察对正立和倒立的威胁性身体表情（恐惧和愤怒）的识别。行为结果发现倒置效应存在。ERP结果发现：早期阶段已有对威胁性情绪（P1）和倒置效应（N1）的加工;中期阶段主要存在对倒置效应的加工（N170）;晚期阶段,威胁性情绪在正立条件上得到更多的加工（P3和LPP）。结果表明,威胁性身体表情加工有独特的时间进程,加工方式以构形加工为主。     

Early Development of Mushroom Bodies in the Brain of the Honeybee Apis mellifera as Revealed by BrdU Incorporation and Ablation Experiments

In the honeybee the mushroom bodies are prominent neuropil structures arranged as pairs in the dorsal protocerebrum of the brain. Each mushroom body is composed of a medial and a lateral subunit. To understand their development, the proliferation pattern of mushroom body intrinsic cells, the Kenyon cells, were examined during larval and pupal stages using the bromodeoxyuridine (BrdU) technique and chemical ablation with hydroxyurea.     

Associative representational plasticity in the auditory cortex: a synthesis of two disciplines

Historically, sensory systems have been largely ignored as potential loci of information storage in the neurobiology of learning and memory. They continued to be relegated to the role of "sensory analyzers" despite consistent findings of associatively induced enhancement of responses in primary sensory cortices to behaviorally important signal stimuli, such as conditioned stimuli (CS), during classical conditioning. This disregard may have been promoted by the fact that the brain was interrogated using only one or two stimuli, e.g., a CS(+) sometimes with a CS(-), providing little insight into the specificity of neural plasticity. This review describes a novel approach that synthesizes the basic experimental designs of the experimental psychology of learning with that of sensory neurophysiology. By probing the brain with a large stimulus set before and after learning, this unified method has revealed that associative processes produce highly specific changes in the receptive fields of cells in the primary auditory cortex (A1). This associative representational plasticity (ARP) selectively facilitates responses to tonal CSs at the expense of other frequencies, producing tuning shifts toward and to the CS and expanded representation of CS frequencies in the tonotopic map of A1. ARPs have the major characteristics of associative memory: They are highly specific, discriminative, rapidly acquired, exhibit consolidation over hours and days, and can be retained indefinitely. Evidence to date suggests that ARPs encode the level of acquired behavioral importance of stimuli. The nucleus basalis cholinergic system is sufficient both for the induction of ARPs and the induction of specific auditory memory. Investigation of ARPs has attracted workers with diverse backgrounds, often resulting in behavioral approaches that yield data that are difficult to interpret. The advantages of studying associative representational plasticity are emphasized, as is the need for greater behavioral sophistication.     

A role for Synapsin in associative learning: the Drosophila larva as a study case

Synapsins are evolutionarily conserved, highly abundant vesicular phosphoproteins in presynaptic terminals. They are thought to regulate the recruitment of synaptic vesicles from the reserve pool to the readily-releasable pool, in particular when vesicle release is to be maintained at high spiking rates. As regulation of transmitter release is a prerequisite for synaptic plasticity, we use the fruit fly Drosophila to ask whether Synapsin has a role in behavioral plasticity as well; in fruit flies, Synapsin is encoded by a single gene (syn). We tackled this question for associative olfactory learning in larval Drosophila by using the deletion mutant syn^97CS, which had been backcrossed to the Canton-S wild-type strain (CS) for 13 generations. We provide a molecular account of the genomic status of syn^97CS by PCR and show the absence of gene product on Western blots and nerve-muscle preparations. We found that olfactory associative learning in syn^97CS larvae is reduced to ~50% of wild-type CS levels; however, responsiveness to the to-be-associated stimuli and motor performance in untrained animals are normal. In addition, we introduce two novel behavioral control procedures to test stimulus responsiveness and motor performance after “sham training.” Wild-type CS and syn^97CS perform indistinguishably also in these tests. Thus, larval Drosophila can be used as a case study for a role of Synapsin in associative learning.     

What is learned in sequential learning? An associative model of reward magnitude serial-pattern learning

A computational model of sequence learning is described that is based on pairwise associations and generalization. Simulations by the model predicted that rats should learn a long monotonic pattern of food quantities better than a nonmonotonic pattern, as predicted by rule-learning theory, and that they should learn a short nonmonotonic pattern with highly discriminable elements better than 1 with less discriminable elements, as predicted by interitem association theory. In 2 other studies, the model also simulated behavioral "rule generalization," "extrapolation," and associative transfer data motivated by both rule-learning and associative perspectives. Although these simulations do not rule out the possibility that rats can use rule induction to learn serial patterns, they show that a simple associative model can account for the classical behavioral studies implicating rule learning in reward magnitude serial-pattern learning.     

Mapping of the Anatomical Circuit of CaM Kinase-Dependent Courtship Conditioning in Drosophila

Globally inhibiting CaM kinase activity in Drosophila, using a variety of genetic techniques, disrupts associative memory yet leaves visual and chemosensory perception intact. These studies implicate CaM kinase in the plastic processes underlying learning and memory but do not identify the neural circuitry that specifies the behavior. In this study, we use the GAL4/UAS binary expression system to define areas of the brain that require CaM kinase for modulation of courtship conditioning. The CaM kinase-dependent neurons that determine the response to the mated female during conditioning and those involved in formation and expression of memory were found to be located in distinct areas of the brain. This supports the idea that courtship conditioning results in two independent behavioral modifications: a decrement in courtship during the conditioning period and an associative memory of conditioning. This study has allowed us for the first time to genetically determine the circuit of information flow for a memory process in Drosophila. The map we have generated dissects the behavior into multiple components and will provide tools that allow both molecular and electrophysiological access to this circuit.     

Immunocytochemical Mapping of an RDL-Like GABA Receptor Subunit and of GABA in Brain Structures Related to Learning and Memory in the Cricket Acheta domesticus

The distribution of putative RDL-like GABA receptors and of γ-aminobutyric acid (GABA) in the brain of the adult house cricket Acheta domesticus was studied using specific antisera. Special attention was given to brain structures known to be related to learning and memory. The main immunostaining for the RDL-like GABA receptor was observed in mushroom bodies, in particular the upper part of mushroom body peduncle and the two arms of the posterior calyx. Weaker immunostaining was detected in the distal part of the peduncle and in the α and β lobes. The dorso- and ventrolateral protocerebrum neuropils appeared rich in RDL-like GABA receptors. Staining was also detected in the glomeruli of the antennal lobe, as well as in the ellipsoid body of the central complex. Many neurons clustered in groups exhibit GABA-like immunoreactivity. Tracts that were strongly immunostained innervated both the calyces and the lobes of mushroom bodies. The glomeruli of the antennal lobe, the ellipsoid body, as well as neuropils of the dorso- and ventrolateral protocerebrum were also rich in GABA-like immuno- reactivity. The data demonstrated a good correlation between the distribution of the GABA-like and of the RDL-like GABA receptor immunoreactivity. The prominent distribution of RDL-like GABA receptor subunits, in particular areas of mushroom bodies and antennal lobes, underlines the importance of inhibitory signals in information processing in these major integrative centers of the insect brain.     

Brain composition and olfactory learning in honey bees

Correlations between brain or brain component size and behavioral measures are frequently studied by comparing different animal species, which sometimes introduces variables that complicate interpretation in terms of brain function. Here, we have analyzed the brain composition of honey bees (Apis mellifera) that have been individually tested in an olfactory learning paradigm. We found that the total brain size correlated with the bees’ learning performance. Among different brain components, only the mushroom body, a structure known to be involved in learning and memory, showed a positive correlation with learning performance. In contrast, visual neuropils were relatively smaller in bees that performed better in the olfactory learning task, suggesting modality-specific behavioral specialization of individual bees. This idea is also supported by inter-individual differences in brain composition. Some slight yet statistically significant differences in the brain composition of European and Africanized honey bees are reported. Larger bees had larger brains, and by comparing brains of different sizes, we report isometric correlations for all brain components except for a small structure, the central body.