Kohei Ueno

Trehalose sensitivity in Drosophila correlates with mutations in and expression of the gustatory receptor gene Gr5a

Drosophila taste gene Tre is located on the distal X chromosome and controls gustatory sensitivity to a subset of sugars [1, 2]. Two adjacent, seven-transmembrane domain genes near the Tre locus are candidate genes for Tre. One (CG3171) encodes a rhodopsin family G protein receptor [3, 4], and the other (Gr5a) is a member of a chemosensory gene family encoding a putative gustatory receptor [5-7]. We carried out molecular analyses of mutations in Tre to elucidate their involvement in the gustatory phenotype. Here, we show that Tre mutations induced by P element-mediated genomic deletions disrupt Gr5a gene organization and the expression of Gr5a mRNA, while disruption of the CG3171 gene or its expression was not always associated with mutations in Tre. In flies with the spontaneous mutation Tre(01), both CG3171 and Gr5a mRNAs are transcribed. Coding sequences of these two candidate genes were compared among various strains. A total of three polymorphic sites leading to amino acid changes in CG3171 were not correlated with the gustatory phenotype. Among four nonsynonymous sites in Gr5a, a single nucleotide polymorphism leading to an Ala218Thr substitution in the predicted second intracellular loop cosegregated with Tre(01). Taken together, the mutation analyses support that Gr5a is allelic to Tre.

Fasting Launches CRTC to Facilitate Long-Term Memory Formation in Drosophila

Hunger and Memory During starvation, are all brain functions slowed down, or are specific functions disabled to save energy? Plaçais and Preat (p. 440) investigated how the brain of Drosophila deals with severe resource limitation. The brain cut selected expenses to reduce the threat to survival and switched off the formation of aversive long-term memory that depends on costly protein synthesis. However, Hirano et al. (p. 443) focused on mild food-deprivation, which actually enhanced long-term memory formation. Presumably, improved memory should enhance survival when competing for limited food. After longer food deprivation, enhancement of aversive long-term memory decreased, while that of appetitive long-term memory remained high: Presumably, as starvation nears, it becomes more important to pursue food at all costs, and so appetitive memory takes precedence over aversive memories. Different types of memory interact in fed versus starved states to promote survival-oriented behavior. Canonical aversive long-term memory (LTM) formation in Drosophila requires multiple spaced trainings, whereas appetitive LTM can be formed after a single training. Appetitive LTM requires fasting prior to training, which increases motivation for food intake. However, we found that fasting facilitated LTM formation in general; aversive LTM formation also occurred after single-cycle training when mild fasting was applied before training. Both fasting-dependent LTM (fLTM) and spaced training–dependent LTM (spLTM) required protein synthesis and cyclic adenosine monophosphate response element–binding protein (CREB) activity. However, spLTM required CREB activity in two neural populations—mushroom body and DAL neurons—whereas fLTM required CREB activity only in mushroom body neurons. fLTM uses the CREB coactivator CRTC, whereas spLTM uses the coactivator CBP. Thus, flies use distinct LTM machinery depending on their hunger state.

Perlecan regulates bidirectional Wnt signaling at the Drosophila neuromuscular junction.

Perlecan/Trol at the neuromuscular junction suppresses presynaptic canonical Wg signaling but enhances the postsynaptic Frizzled nuclear import pathway.

Glial Dysfunction Causes Age-Related Memory Impairment in Drosophila

Several aging phenotypes, including age-related memory impairment (AMI), are thought to be caused by cumulative oxidative damage. In Drosophila, age-related impairments in 1 hr memory can be suppressed by reducing activity of protein kinase A (PKA). However, the mechanism for this effect has been unclear. Here we show that decreasing PKA suppresses AMI by reducing activity of pyruvate carboxylase (PC), a glial metabolic enzyme whose amounts increase upon aging. Increased PC activity causes AMI through a mechanism independent of oxidative damage. Instead, increased PC activity is associated with decreases in D-serine, a glia-derived neuromodulator that regulates NMDA receptor activity. D-serine feeding suppresses both AMI and memory impairment caused by glial overexpression of dPC, indicating that an oxidative stress-independent dysregulation of glial modulation of neuronal activity contributes to AMI in Drosophila.

Adenylyl cyclase encoded by AC78C participates in sugar perception in Drosophila melanogaster

In gustatory receptor neurons (GRNs) in Drosophila melanogaster, Gr5a and one of the Gr64s encode sugar receptors with seven transmembrane domains. Previously, we have shown that the responses to various sugars are depressed in DGsα mutant flies ( Ueno et al., 2006 ). Because DGsα is a homolog of Gs, we hypothesized that the sugar receptors are coupled to adenylyl cyclase (AC) in Drosophila. The aim of this study was to identify the AC that participates in sugar perception. Here, we found that an AC inhibitor, MDL‐12330A, depressed the response in GRNs to trehalose as well as sucrose; that an AC gene, AC78C, was expressed in the sugar‐sensitive GRNs; that RNAi against AC78C depressed the electrical response in GRNs to sucrose; and that the sugar response in GRNs, as well as sugar intake in a behavioral assay in an AC78C mutant, was depressed at low sugar concentrations. We conclude that AC78C, via cAMP, participates in the sugar‐taste signaling pathway at the low concentration range.

A putative binding protein for lipophilic substances related to butterfly oviposition

A unique protein of 23 kDa (Jf23) was found in the tarsus of the female swallowtail butterfly, Atrophaneura alcinous. Jf23 has 38% identity with a bilin‐binding protein, which was found in the cabbage butterfly, Pieris brassicae, and which has two consensus sequences in common with the members of the lipocalin family, suggesting that it is a binding protein for lipophilic ligands. Western blot analysis showed that Jf23 was expressed only in the female, and not in the male. Electrophysiological response of the female tarsi was stimulated by methanolic extract of their host plant, Dutchmans pipe (Aristolochia debilis). The stimulated response was depressed by the presence of Jf23 antiserum. These results suggest that Jf23 is one of the chemosensory signaling proteins, which plays one or more roles in female butterfly oviposition.

Long‐term enhancement of synaptic transmission between antennal lobe and mushroom body in cultured Drosophila brain

•  During olfactory aversive conditioning in Drosophila, odour and shock information are delivered to the mushroom bodies (MBs) through projection neurons in the antennal lobes (ALs) and ascending fibres of the ventral nerve cord (AFV), respectively. •  Using an isolated cultured brain expressing a Ca2+ indicator in the MBs, we demonstrated that the simultaneous stimulation of the ALs and AFV establishes long‐term enhancement (LTE) in AL‐induced Ca2+ responses. •  The physiological properties of LTE, including associativity, input specificity and persistence, are highly reminiscent of those of olfactory memory. •  Similar to olfactory aversive memory, LTE requires the activation of nicotinic acetylcholine receptors that mediate the AL‐evoked Ca2+ response, NMDA receptors that mediate the AFV‐induced Ca2+ response, and D1 dopamine receptors during the simultaneous stimulation of the ALs and AFV. •  Considering the physiological and genetic analogies, we propose that LTE at the AL–MB synapse can be a relevant cellular model for olfactory memory.

Two types of Ca2+ channel linked to two endocytic pathways coordinately maintain synaptic transmission at the Drosophila synapse

Endocytosis at the presynaptic terminal is initiated by Ca2+ influx through voltage‐gated Ca2+ channels. At the Drosophila neuromuscular junction, we demonstrated two components of endocytosis linked to distinct Ca2+ channels. A voltage‐gated Ca2+ channel blocker, (R)‐(+)‐Bay K8644 (R‐BayK), selectively blocked one component (R‐BayK‐sensitive component) without affecting exocytosis, while low concentrations of La3+ preferentially depressed the other component (La3+ ‐sensitive component). In a temperature‐sensitive mutant, shibirets, at non‐permissive temperatures, dynamin clusters were found immunohistochemically at the active zone (AZ) during the R‐BayK‐sensitive endocytosis, while they were detected at the non‐AZ during the La3+‐sensitive endocytosis. Immunostaining of the Ca2+ channel α2δ subunit encoded by straightjacket (stj) was found within the AZ, and a mutation in stj depressed the R‐BayK‐sensitive component but enhanced the La3+ ‐sensitive one, indicating that the α2δ subunit is associated with the R‐BayK‐sensitive Ca2+ channel. Filipin bound to the non‐AZ membrane and inhibited the La3+ ‐sensitive component, but not the R‐BayK‐sensitive one. We concluded that the R‐BayK‐sensitive component of endocytosis occurred at the AZ and termed this AZ endocytosis. We also concluded that the La3+ ‐sensitive component occurred at the non‐AZ and termed this non‐AZ endocytosis. These two types of endocytosis were modulated by various drugs towards opposite directions, indicating that they were differentially regulated. During high‐frequency stimulation, AZ endocytosis operated mainly in the early phase, whereas non‐AZ endocytosis operated in the late phase. Thus, intense synaptic transmission is coordinately maintained by synaptic vesicle recycling initiated by Ca2+ influx through the two types of Ca2+ channel.

Coincident postsynaptic activity gates presynaptic dopamine release to induce plasticity in Drosophila mushroom bodies

Simultaneous stimulation of the antennal lobes (ALs) and the ascending fibers of the ventral nerve cord (AFV), two sensory inputs to the mushroom bodies (MBs), induces long-term enhancement (LTE) of subsequent AL-evoked MB responses. LTE induction requires activation of at least three signaling pathways to the MBs, mediated by nicotinic acetylcholine receptors (nAChRs), NMDA receptors (NRs), and D1 dopamine receptors (D1Rs). Here, we demonstrate that inputs from the AL are transmitted to the MBs through nAChRs, and inputs from the AFV are transmitted by NRs. Dopamine signaling occurs downstream of both nAChR and NR activation, and requires simultaneous stimulation of both pathways. Dopamine release requires the activity of the rutabaga adenylyl cyclase in postsynaptic MB neurons, and release is restricted to MB neurons that receive coincident stimulation. Our results indicate that postsynaptic activity can gate presynaptic dopamine release to regulate plasticity. DOI: http://dx.doi.org/10.7554/eLife.21076.001

Learning defects in Drosophila growth restricted chico mutants are caused by attenuated adenylyl cyclase activity

BackgroundReduced insulin/insulin-like growth factor signaling (IIS) is a major cause of symmetrical intrauterine growth retardation (IUGR), an impairment in cell proliferation during prenatal development that results in global growth defects and mental retardation. In Drosophila, chico encodes the only insulin receptor substrate. Similar to other animal models of IUGR, chico mutants have defects in global growth and associative learning. However, the physiological and molecular bases of learning defects caused by chico mutations, and by symmetrical IUGR, are not clear.ResultsIn this study, we found that chico mutations impair memory-associated synaptic plasticity in the mushroom bodies (MBs), neural centers for olfactory learning. Mutations in chico reduce expression of the rutabaga-type adenylyl cyclase (rut), leading to decreased cAMP synthesis in the MBs. Expressing a rut+ transgene in the MBs restores memory-associated plasticity and olfactory associative learning in chico mutants, without affecting growth. Thus chico mutations disrupt olfactory learning, at least in part, by reducing cAMP signaling in the MBs.ConclusionsOur results suggest that some cognitive defects associated with reduced IIS may occur, independently of developmental defects, from acute reductions in cAMP signaling.