TY - JOUR
T1 - Electrophysiological method for whole-cell voltage clamp recordings from drosophila photoreceptors
AU - Katz, Ben
AU - Gutorov, Rita
AU - Rhodes-Mordov, Elisheva
AU - Hardie, Roger C.
AU - Minke, Baruch
N1 - Publisher Copyright:
© 2017 Journal of Visualized Experiments.
PY - 2017/6/13
Y1 - 2017/6/13
N2 - Whole-cell voltage clamp recordings from Drosophila melanogaster photoreceptors have revolutionized the field of invertebrate visual transduction, enabling the use of D. melanogaster molecular genetics to study inositol-lipid signaling and Transient Receptor Potential (TRP) channels at the single-molecule level. A handful of labs have mastered this powerful technique, which enables the analysis of the physiological responses to light under highly controlled conditions. This technique allows control over the intracellular and extracellular media; the membrane voltage; and the fast application of pharmacological compounds, such as a variety of ionic or pH indicators, to the intra-and extracellular media. With an exceptionally high signal-to-noise ratio, this method enables the measurement of dark spontaneous and light-induced unitary currents (i.e. spontaneous and quantum bumps) and macroscopic Light-induced Currents (LIC) from single D. melanogaster photoreceptors. This protocol outlines, in great detail, all the key steps necessary to perform this technique, which includes both electrophysiological and optical recordings. The fly retina dissection procedure for the attainment of intact and viable ex vivo isolated ommatidia in the bath chamber is described. The equipment needed to perform whole-cell and fluorescence imaging measurements are also detailed. Finally, the pitfalls in using this delicate preparation during extended experiments are explained.
AB - Whole-cell voltage clamp recordings from Drosophila melanogaster photoreceptors have revolutionized the field of invertebrate visual transduction, enabling the use of D. melanogaster molecular genetics to study inositol-lipid signaling and Transient Receptor Potential (TRP) channels at the single-molecule level. A handful of labs have mastered this powerful technique, which enables the analysis of the physiological responses to light under highly controlled conditions. This technique allows control over the intracellular and extracellular media; the membrane voltage; and the fast application of pharmacological compounds, such as a variety of ionic or pH indicators, to the intra-and extracellular media. With an exceptionally high signal-to-noise ratio, this method enables the measurement of dark spontaneous and light-induced unitary currents (i.e. spontaneous and quantum bumps) and macroscopic Light-induced Currents (LIC) from single D. melanogaster photoreceptors. This protocol outlines, in great detail, all the key steps necessary to perform this technique, which includes both electrophysiological and optical recordings. The fly retina dissection procedure for the attainment of intact and viable ex vivo isolated ommatidia in the bath chamber is described. The equipment needed to perform whole-cell and fluorescence imaging measurements are also detailed. Finally, the pitfalls in using this delicate preparation during extended experiments are explained.
KW - Current clamp
KW - Drosophila melanogaster
KW - Electrophysiology
KW - Intracellular perfusion
KW - Issue 124
KW - Neuroscience
KW - Phototransduction
KW - Single-cell Ca imaging
KW - Voltage clamp
KW - Whole-cell recordings
UR - http://www.scopus.com/inward/record.url?scp=85021195463&partnerID=8YFLogxK
U2 - 10.3791/55627
DO - 10.3791/55627
M3 - ???researchoutput.researchoutputtypes.contributiontojournal.article???
C2 - 28654039
AN - SCOPUS:85021195463
SN - 1940-087X
VL - 2017
JO - Journal of Visualized Experiments
JF - Journal of Visualized Experiments
IS - 124
M1 - e55627
ER -