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Full Abstracts – NEUROPHYSIOLOGY AND BEHAVIOR
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stallone
and
balboa
are DEG/ENaC genes required for mechanical nociception.
Stephanie Mauthner
1
, Richard Hwang
2
, Jason Caldwell
3
, W. Daniel
Tracey
1,2,3
. 1) Univ Prog in Genetics and Genomics; 2) Dept of Neurobiology; 3) Dept of Anesthesiology, Duke University, Durham, NC.
Drosophila
larvae respond to potentially tissue-damaging stimuli with nocifensive escape locomotion (NEL). NEL is a stereotyped withdrawal behavior in
which the larva rotates in a “corkscrew” pattern distinct from normal locomotion and is triggered by noxious heat (>39oC) or noxious mechanical (>30mN)
stimuli. The class IV multidendritic (md) neurons are the polymodal nociceptors responsible for triggering NEL. Recent evidence suggests that the
pickpocket
(
ppk
) gene, a degenerin/epithelial sodium channel (Deg/ENaC) subunit, is involved in the mechanotransduction of these neurons.
ppk
mutants
show reduced NEL responses to noxious mechanical stimuli without showing defects in thermal or optogenetic NEL responses. While removal of this ion
channel diminishes the larval response to noxious mechanical force, it does not completely abolish it leading us to hypothesize that additional
mechanonociceptive channels have yet to be identified. The objective of this study was to 1) identify novel channels and 2) functionally characterize their
role in mechanical nociception. We carried out an
in vivo
forward genetic screen by utilizing the VDRC RNAi collection to reduce expression of all known
and predicted ion channels specifically in the class IV md nociceptors. Taking this approach, two novel genes showing a reduced mechanical response were
identified. We named these genes
stallone
and
balboa
. Remarkably, both genes are predicted to encode Deg/ENaC subunits suggesting an important role for
this ion channel family in sensory perception of noxious stimuli. In addition, neither gene is required for optogenetic NEL responses indicating that both
stallone
and
balboa
function at or near the transduction step, perhaps forming a multimeric channel with ppk. We used a hybrid element insertion (HEI)
approach to delete the stallone locus; this genetic mutant phenocopies mechanical nociception defects observed in RNAi mutants.
57
Analysis of escape and avoidance behavior in
Drosophila
larvae.
Tomoko Ohyama, James Truman, Rex Kerr, Marta Zlatic. Janelia Farm Research
Campus/HHMI, Ashburn, VA.
Nervous systems, which allow organisms to respond flexibly to their environments, must transform the sensory inputs they receive into appropriate
behavioral outputs. To study this we use the somatosensory system of
Drosophila
larvae. All somatosensory neurons have been anatomically identified and
are known to project to the ventral nerve cord (VNC). The VNC contains a relatively small number of neuronal classes, and the Truman lab has identified a
GAL4 line for each of these (generated by the Rubin lab). These Gal4 lines allow us to test the function of each class of interneuron downstream of
somatosensory circuits. We have established a high-throughput behavioral analysis system to elucidate the function of each neuron class. We use a tracking
software developed by Kerr Lab for
C. elegans
, to monitor the reactions of larvae. This system allows us to analyze the dynamic parameters (speed,
curvature, hunching, change of direction, etc.) of populations of animals as well as of individual animals. First, we analyzed the reactions to vibration and
pain. We found that larvae show stereotypical reaction sequences during continuous vibration—stop (startle) > head retraction > turning > forward crawling
(avoidance) > off response—and pain stimulation—roll > rapid forward crawling. By inactivating sensory neurons, we also confirmed that chordotonal (ch)
neurons are necessary for sensing vibration and that multi-dendritic type IV (MD IV) neurons are necessary for sensing pain-inducing stimuli. Similar but
different motor patterns (avoidance and escape; fast forward crawling) elicited by vibration and pain-inducing stimuli pose an interesting question as to how
the circuitries downstream of ch and MD IV neurons elicit similar but non-identical reaction sequences. We performed inactivating/activating each
interneuron class in the VNC to assess the correspondence between sensorimotor processing and neural connectivity in the circuits associated with ch and
MD IV neurons. We found several interneuron classes that involed in vibration and pain stimulations.
58
Decision-making neurons for feeding behavior revealed by genetic activation in
Drosophila
.
Motojiro Yoshihara
1
, Thomas Flood
1
, Michael Gorczyca
1
,
Shinya Iguchi
1
, Benjamin White
2
, Kei Ito
3
. 1) Neurobiology, UMass Medical School, Worcester, MA; 2) Laboratory of Molecular Biology, NIMH,
Bethesda, MD; 3) Institute of Molecular and Cellular Biosciences, University of Tokyo, Tokyo, Japan.
The decision of when to eat is a complex function of both environmental variables and internal physiological state. How these external and internal
determinants are integrated by the nervous system is largely unknown and the neural substrates of the feeding decision remain poorly characterized. We
randomly activated subsets of the
Drosophila
brain in 835 Gal 4 lines established by the NP consortium
(1)
through activation of a cold-activated channel,
TRPM8
(2)
, or a heat-activated channel, TrpA1
(3)
in Gal 4 expressing cells. In the unbiased screening, we identified a Gal 4 line showing feeding behavior. By
restricting the Gal 4 expression through the “flip-out Gal80” technique, we identified a critical pair of neurons, Fdg (feeding) neurons, in the brain that
induced the entire feeding sequence when activated. The large dendritic arbor of the Fdg-neurons suggests a role in integrating multiple information types.
Consistent with this, functional calcium imaging revealed that Fdg-neurons responded to food presentation only in the starved state. The study of Fdg-neuron
function may help elucidate how feeding decisions are made generally. 1) Yoshihara and Ito (2000) Dros. Inf. Ser. 83:199. 2) Peabody et al. (2009) J
Neurosci 29, 3343-3353. 3) Hamada et al. (2008) Nature 454, 217-220.
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A long-term memory circuit from mushroom bodies to central complex in the Drosophila brain.
Tsung-Pin Pai, Ann-Shyn Chiang. Institute of
Biotechnology/Brain Research Center, National Tsing Hua University, Hsinchu 30013, Taiwan.
A functional memory circuit must (i) register (acquire) an experience via a persistent neural activity, (ii) consolidate (store) a lasting memory via (protein
synthesis-dependent) structural/functional changes somewhere in that circuit and (iii) retrieve a long-term memory via reactivation of (some or all) of the
circuit. Neural activity in the mushroom body (MB) neurons contributes to acquisition, consolidation and retrieval of olfactory associative long-term
memory (LTM). We found that outputs of speicific MB effernt neurons giving dendrites in the verticle lobes are required for LTM retrieval. These neurons
were intrinsically responsive to a diversity of odors and exhibited enhanced GCaMP activity to the conditioned odors. Using GFP Reconstitution Across
Synaptic Partners (GRASP) labeling, we identified a group of neurons interconnected between MB effernt neurons and the fan-shape body (FB), the
premotor center controlling Drosophila locomotion. A behavioral screen for neurons in which their neurotransmission outputs are required for LTM retrieval
is still ongoing. We will present a comprehansive neural circuit in which neurons are structurally and functionally interconnected for LTM retrieval.