Targeted Attenuation of Electrical Activity in Drosophila Using a Genetically Modified K+ Channel

White, Benjamin H.; Osterwalder, Thomas; Yoon, Kenneth S.; Joiner, William J.; Whim, Matthew D.; Kaczmarek, Leonard K.; Keshishian, Haig

doi:10.1016/s0896-6273(01)00415-9

Cited by 130 publications

(151 citation statements)

References 45 publications

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“…The w,rpr; ϩ;ϩ, w and Canton-S lines were from the Bloomington Stock Center (Indiana University, Bloomington, IN). The 1ϫ, 2ϫ, and 3ϫ electrical knock-out (EKO) lines have been described previously (White et al, 2001).…”

Section: Methodsmentioning

confidence: 99%

“…Previously, we have demonstrated the efficacy of targeted suppression of neuronal excitability in analyzing neuronal function (White et al, 2001;Nitabach et al, 2002). Applying this approach, we suppressed membrane excitability in N CCAP to ask whether hormone secretion requires electrical activity in these neurons.…”

Section: Suppression Of Excitability In N Ccap Inhibits Wing Expansiomentioning

confidence: 99%

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Functional Dissection of a Neuronal Network Required for Cuticle Tanning and Wing Expansion inDrosophila

Luan¹,

Lemon²,

Peabody³

et al. 2006

J. Neurosci.

177

240

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A subset of Drosophila neurons that expresses crustacean cardioactive peptide (CCAP) has been shown previously to make the hormone bursicon, which is required for cuticle tanning and wing expansion after eclosion. Here we present evidence that CCAP-expressing neurons (N CCAP ) consist of two functionally distinct groups, one of which releases bursicon into the hemolymph and the other of which regulates its release. The first group, which we call N CCAP -c929, includes 14 bursicon-expressing neurons of the abdominal ganglion that lie within the expression pattern of the enhancer-trap line c929-Gal4. We show that suppression of activity within this group blocks bursicon release into the hemolymph together with tanning and wing expansion. The second group, which we call N CCAP -R, consists of N CCAP neurons outside the c929-Gal4 pattern. Because suppression of synaptic transmission and protein kinase A (PKA) activity throughout N CCAP , but not in N CCAP -c929, also blocks tanning and wing expansion, we conclude that neurotransmission and PKA are required in N CCAP -R to regulate bursicon secretion from N CCAP -c929. Enhancement of electrical activity in N CCAP -R by expression of the bacterial sodium channel NaChBac also blocks tanning and wing expansion and leads to depletion of bursicon from central processes. NaChBac expression in N CCAP -c929 is without effect, suggesting that the abdominal bursicon-secreting neurons are likely to be silent until stimulated to release the hormone. Our results suggest that N CCAP form an interacting neuronal network responsible for the regulation and release of bursicon and suggest a model in which PKA-mediated stimulation of inputs to normally quiescent bursicon-expressing neurons activates release of the hormone.

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Section: Methodsmentioning

confidence: 99%

Section: Suppression Of Excitability In N Ccap Inhibits Wing Expansiomentioning

confidence: 99%

Functional Dissection of a Neuronal Network Required for Cuticle Tanning and Wing Expansion inDrosophila

Luan¹,

Lemon²,

Peabody³

et al. 2006

J. Neurosci.

177

240

View full text Add to dashboard Cite

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“…Molecular Biology. SDN was generated from the EKO construct (32). The UAS-EKO ϩ vector was cleaved at the end of the ORF and 3Ј to the coding region for the S1 transmembrane helix.…”

Section: Methodsmentioning

confidence: 99%

“…Larval K ϩ currents were examined by using a two electrode voltage clamp similar to above. Larvae were dissected in 0 mM Ca 2ϩ physiological saline with 0.5 mM EGTA to eliminate the I CF and I CS calcium currents (32). The muscle was clamped at Ϫ80 mV and stepped to ϩ30 mV in 10-mV increments.…”

Section: Methodsmentioning

confidence: 99%

Dissection of synaptic excitability phenotypes by using a dominant-negative Shaker K ⁺ channel subunit

Mosca

Carrillo

White

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

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During nervous system development, synapses undergo morphological change as a function of electrical activity. In Drosophila, enhanced activity results in the expansion of larval neuromuscular junctions. We have examined whether these structural changes involve the pre-or postsynaptic partner by selectively enhancing electrical excitability with a Shaker dominant-negative (SDN) potassium channel subunit. We find that the SDN enhances neurotransmitter release when expressed in motoneurons, postsynaptic potential broadening when expressed in muscles and neurons, and selectively suppresses fast-inactivating, Shaker-mediated I A currents in muscles. SDN expression also phenocopies the canonical behavioral phenotypes of the Sh mutation. At the neuromuscular junction, we find that activity-dependent changes in arbor size occur only when SDN is expressed presynaptically. This finding indicates that elevated postsynaptic membrane excitability is by itself insufficient to enhance presynaptic arbor growth. Such changes must minimally involve increased neuronal excitability.activity ͉ behavioral genetics ͉ Drosophila E lectrical activity plays a prominent role in the development and plasticity of synapses (1-3). The glutamatergic neuromuscular junction (NMJ) of Drosophila is favorable for examining synaptogenesis and the role of electrical activity in synaptic growth and refinement (4, 5). In Drosophila, activity influences synaptic connectivity (6), size (7-9), and homeostasis (10, 11). Activity may also regulate retrograde signals that influence NMJ development (12)(13)(14). However, determining the relative contributions of pre-and postsynaptic excitability to synaptic development remains a significant challenge.Addressing this problem requires molecular tools that selectively alter excitability on either side of the synapse. Early success in suppressing excitability has involved the targeted expression of modified ion channels or receptors (10,15,16). A promising approach for enhancing electrical activity involves the dominantnegative suppression of K ϩ currents involved in membrane repolarization and excitability (17, 18).The Shaker (Sh) type I A K ϩ current of Drosophila plays a key role in regulating membrane excitability of neurons and muscles (19,20) and also influences the processing of graded potentials (21). Sh mutants have well characterized hyperactive behavioral and electrophysiological phenotypes (20,(22)(23)(24), making Sh a good candidate for dominant-negative suppression. Hyperexcitable Sh mutants, when combined with other K ϩ channel mutants such as ether-a-go-go, also have enlarged larval NMJs (7-9). However, whether these structural changes arise from pre-or postsynaptic membrane hyperexcitability has remained unresolved, because the Sh mutation affects both neurons and muscle.To dissect these changes, we have developed a Sh dominantnegative (SDN) transgene as a tool for targeted enhancement of electrical excitability. SDN effectively suppresses I A, as demonstrated by voltage-clamp analysis of muscle...

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“…Recently, it was demonstrated that neuronal circuits can be manipulated by expressing mutated ion channels or G protein-coupled receptors (GPCRs). For example, the regional expression of a genetically modified K ϩ channel in Drosophila was able to reduce the excitability of targeted cells (i.e., muscle, neurons, photoreceptors) (1). Silencing of cortical neurons was achieved by binding of the peptide allostatin to its exogenously expressed receptor (2).…”

mentioning

confidence: 99%

Fast noninvasive activation and inhibition of neural and network activity by vertebrate rhodopsin and green algae channelrhodopsin

Gutierrez

Hanson

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

493

420

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Techniques for fast noninvasive control of neuronal excitability will be of major importance for analyzing and understanding neuronal networks and animal behavior. To develop these tools we demonstrated that two light-activated signaling proteins, vertebrate rat rhodopsin 4 (RO4) and the green algae channelrhodospin 2 (ChR2), could be used to control neuronal excitability and modulate synaptic transmission. Vertebrate rhodopsin couples to the Gi͞o, pertussis toxin-sensitive pathway to allow modulation of G protein-gated inward rectifying potassium channels and voltagegated Ca 2؉ channels. Light-mediated activation of RO4 in cultured hippocampal neurons reduces neuronal firing within ms by hyperpolarization of the somato-dendritic membrane and when activated at presynaptic sites modulates synaptic transmission and paired-pulse facilitation. In contrast, somato-dendritic activation of ChR2 depolarizes neurons sufficiently to induce immediate action potentials, which precisely follow the ChR2 activation up to light stimulation frequencies of 20 Hz. To demonstrate that these constructs are useful for regulating network behavior in intact organisms, embryonic chick spinal cords were electroporated with either construct, allowing the frequency of episodes of spontaneous bursting activity, known to be important for motor circuit formation, to be precisely controlled. Thus light-activated vertebrate RO4 and green algae ChR2 allow the antagonistic control of neuronal function within ms to s in a precise, reversible, and noninvasive manner in cultured neurons and intact vertebrate spinal cords.A major challenge in understanding the relationship between neural activity and development and between neuronal circuit activity and specific behaviors is to be able to control the activity of large populations of neurons or regions of individual nerve cells simultaneously. Recently, it was demonstrated that neuronal circuits can be manipulated by expressing mutated ion channels or G protein-coupled receptors (GPCRs). For example, the regional expression of a genetically modified K ϩ channel in Drosophila was able to reduce the excitability of targeted cells (i.e., muscle, neurons, photoreceptors) (1). Silencing of cortical neurons was achieved by binding of the peptide allostatin to its exogenously expressed receptor (2). Recently, Zemelman et al. (3) elegantly demonstrated that light activation of the protein complex, encoded by the Drosophila photoreceptor genes (i.e., arrestin-2, rhodopsin, and G protein ␣ subunit), could induce action potential firing of hippocampal neurons. Activation and deactivation of neuronal firing could also be achieved when ligand-gated ion channels, such as the capsaicin receptor, menthol receptor, purinergic receptors, or lightcontrollable K ϩ channel blockers, were used to control firing in hippocampal neurons (4, 5). However, the application of these techniques to control neuronal function especially in neural circuits and living animals is limited by their relatively slow time course, the complex...

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Targeted Attenuation of Electrical Activity in Drosophila Using a Genetically Modified K+ Channel

Cited by 130 publications

References 45 publications

Functional Dissection of a Neuronal Network Required for Cuticle Tanning and Wing Expansion inDrosophila

Functional Dissection of a Neuronal Network Required for Cuticle Tanning and Wing Expansion inDrosophila

Dissection of synaptic excitability phenotypes by using a dominant-negative Shaker K ⁺ channel subunit

Fast noninvasive activation and inhibition of neural and network activity by vertebrate rhodopsin and green algae channelrhodopsin

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Targeted Attenuation of Electrical Activity in Drosophila Using a Genetically Modified K+ Channel

Cited by 130 publications

References 45 publications

Functional Dissection of a Neuronal Network Required for Cuticle Tanning and Wing Expansion inDrosophila

Functional Dissection of a Neuronal Network Required for Cuticle Tanning and Wing Expansion inDrosophila

Dissection of synaptic excitability phenotypes by using a dominant-negative Shaker K + channel subunit

Fast noninvasive activation and inhibition of neural and network activity by vertebrate rhodopsin and green algae channelrhodopsin

Contact Info

Product

Resources

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Dissection of synaptic excitability phenotypes by using a dominant-negative Shaker K ⁺ channel subunit