Abstract
Electrophysiological studies provide essential clues about the regulation and physiological function of ion channel proteins. Probing ion channel activity in vivo, though, often is challenging. This can limit the usefulness of such model organisms as Drosophila for electrophysiological studies. This is unfortunate because these genetically tractable organisms represent powerful research tools that facilitate elaboration of complex questions of physiology. Here, we describe a recently developed method for recording ion channel activity in Drosophila sensory neurons. This approach is based on patch-clamping primary neuron cultures from Drosophila embryos. Such cultures allow the study of ion channels in different genetic backgrounds. In addition to describing how to prepare a primary neuronal cell culture from Drosophila embryos, we discuss, as an example of utility, analysis of Na+ currents in cultured class IV multidendritic (md) sensory neurons with the patch clamp technique. Excitability of md sensory neurons, manifested as action potential firing, is revealed with whole-cell current-clamping. Voltage-clamping class IV md neurons revealed the activity of the voltage-gated Na+ channel, paralytic. Moreover, challenging class IV md neurons with acidic pH activates acid-sensing inward Na+ currents. Genetic manipulation of Drosophila combined with this electrophysiological readout of activity identifies pickpocket1 (Ppk1), a member of the Deg/ENaC channel family, as responsible for conducting an acid-sensing Na+ current in class IV md sensory neurons.
| Original language | English (US) |
|---|---|
| Title of host publication | Methods in Molecular Biology |
| Pages | 385-397 |
| Number of pages | 13 |
| Volume | 998 |
| DOIs | |
| State | Published - 2013 |
Publication series
| Name | Methods in Molecular Biology |
|---|---|
| Volume | 998 |
| ISSN (Print) | 10643745 |
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Keywords
- Drosophila sensory neurons
- Ion channel
- Patch clamping
- Primary cell culture
ASJC Scopus subject areas
- Molecular Biology
- Genetics
Cite this
Patch-clamping drosophila sensory neurons. / Kucher, Volodymyr; Eaton, Benjamin A; Stockand, James D; Boiko, Nina.
Methods in Molecular Biology. Vol. 998 2013. p. 385-397 (Methods in Molecular Biology; Vol. 998).Research output: Chapter in Book/Report/Conference proceeding › Chapter
}
TY - CHAP
T1 - Patch-clamping drosophila sensory neurons
AU - Kucher, Volodymyr
AU - Eaton, Benjamin A
AU - Stockand, James D
AU - Boiko, Nina
PY - 2013
Y1 - 2013
N2 - Electrophysiological studies provide essential clues about the regulation and physiological function of ion channel proteins. Probing ion channel activity in vivo, though, often is challenging. This can limit the usefulness of such model organisms as Drosophila for electrophysiological studies. This is unfortunate because these genetically tractable organisms represent powerful research tools that facilitate elaboration of complex questions of physiology. Here, we describe a recently developed method for recording ion channel activity in Drosophila sensory neurons. This approach is based on patch-clamping primary neuron cultures from Drosophila embryos. Such cultures allow the study of ion channels in different genetic backgrounds. In addition to describing how to prepare a primary neuronal cell culture from Drosophila embryos, we discuss, as an example of utility, analysis of Na+ currents in cultured class IV multidendritic (md) sensory neurons with the patch clamp technique. Excitability of md sensory neurons, manifested as action potential firing, is revealed with whole-cell current-clamping. Voltage-clamping class IV md neurons revealed the activity of the voltage-gated Na+ channel, paralytic. Moreover, challenging class IV md neurons with acidic pH activates acid-sensing inward Na+ currents. Genetic manipulation of Drosophila combined with this electrophysiological readout of activity identifies pickpocket1 (Ppk1), a member of the Deg/ENaC channel family, as responsible for conducting an acid-sensing Na+ current in class IV md sensory neurons.
AB - Electrophysiological studies provide essential clues about the regulation and physiological function of ion channel proteins. Probing ion channel activity in vivo, though, often is challenging. This can limit the usefulness of such model organisms as Drosophila for electrophysiological studies. This is unfortunate because these genetically tractable organisms represent powerful research tools that facilitate elaboration of complex questions of physiology. Here, we describe a recently developed method for recording ion channel activity in Drosophila sensory neurons. This approach is based on patch-clamping primary neuron cultures from Drosophila embryos. Such cultures allow the study of ion channels in different genetic backgrounds. In addition to describing how to prepare a primary neuronal cell culture from Drosophila embryos, we discuss, as an example of utility, analysis of Na+ currents in cultured class IV multidendritic (md) sensory neurons with the patch clamp technique. Excitability of md sensory neurons, manifested as action potential firing, is revealed with whole-cell current-clamping. Voltage-clamping class IV md neurons revealed the activity of the voltage-gated Na+ channel, paralytic. Moreover, challenging class IV md neurons with acidic pH activates acid-sensing inward Na+ currents. Genetic manipulation of Drosophila combined with this electrophysiological readout of activity identifies pickpocket1 (Ppk1), a member of the Deg/ENaC channel family, as responsible for conducting an acid-sensing Na+ current in class IV md sensory neurons.
KW - Drosophila sensory neurons
KW - Ion channel
KW - Patch clamping
KW - Primary cell culture
UR - http://www.scopus.com/inward/record.url?scp=84877073648&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84877073648&partnerID=8YFLogxK
U2 - 10.1007/978-1-62703-351-0_30
DO - 10.1007/978-1-62703-351-0_30
M3 - Chapter
SN - 9781627033503
VL - 998
T3 - Methods in Molecular Biology
SP - 385
EP - 397
BT - Methods in Molecular Biology
ER -