Electrophysiological Recordings from Drosophila Sensilla in Response to Volatile Odorants

Samenvatting

Source: Ng, R., et al. Electrophysiological recording from Drosophila Trichoid sensilla in response to odorants of low volatility. J. Vis. Exp. (2017).

In this video, a live Drosophila fly is mounted in a tube for electrophysiological studies. Electrodes are positioned at the base of the sensillum to record specific neuronal responses to pheromones with the fly's olfactory receptor neurons.

Protocol

1. Preparation of the Hardware for at4 Recording

1. Use a pipette puller instrument to prepare electrodes with aluminosilicate glass capillaries (O.D 1.0 mm, I.D. 0.64 mm). Blunt the tip of the reference electrode slightly with a pair of fine forceps to facilitate insertion into the clypeus of the fly (i.e. a rounded plate at the front of the fly head, above the mouthparts).
   NOTE: 7-day-old WT(Wild type) males (Berlin) were used in this study. Use AHL saline solution as the electrolyte for both electrodes.
2. Prepare 1 L of AHL(Adult Hemolymph-Like) by mixing 900 mL of distilled water with 6.312 g of NaCl, 0.373 g of KC....

Resultaten

Figure 1: Cartridge and Olfactometer Setup. (A) Preparation of odor cartridges. From left to right: a standard 200-µL pipette tip, the first and second cartridge sections, and a completed odorant cartridge. (B) The cartridge connected to the olfactometer, showing .......

Materialen

Lijst van materialen gebruikt in dit artikel

Naam	Bedrijf	Catalogusnummer	Opmerkingen
Prep Setup & Miscellaneous Materials
Pipette Puller Instrument	Sutter Instruments ,Novato CA USA	P97	Pipette Puller
Borosilicate Glass Capillaries	World Precision Instruments, Sarasota FL USA	1B100F-4	to make holding rods
Aluminosilicate Glass Capillaries	Sutter Instruments, Novato CA USA	AF100-64-10	to make electrodes
Superfrost Microscope Slides	Fisher Scientific, Pittsburgh PA USA	12-550-143	for fly-prep station
Permanent Double Sided Tape	Scotch St. Paul MN USA	NA	for fly-prep station
Upright microscope	Olympus, Shinjuku Tokyo Japan	BX51	for recording rig
Plastalina modeling clay	Van Aken, North Charleston SC USA	B0019QZMQQ	for prep station and to stablize the holding rod
Rapid-Flow Sterile Disposable Filter Unit with SFCA Membrane, 0.45 mm	Nalgene, Rochester NY USA	#156-4045	to sterilize AHL solution
200 µL pipette tip	VWR Radnor, PA USA	53508-810	to make odor cartridges and fly prep
Filter Paper	Whatman, Maidstone Kent UK	740-E	to make odor cartridges
Vacuum Desiccator	Cole-Parmer, Vernon Hills IL USA	VX-06514-30	to vaporize ethanol solvent
cis-palmitoleic acid	Cayman Chemical, Ann Arbor MI USA	#10009871 (CAS # 373-49-9)	Or47b odorant
trans-palmitoleic acid	Cayman Chemical, Ann Arbor MI USA	#9001798 (CAS # 10030-73-6)	Or47b odorant
Ethanol	Spectrum Chemical MFG. New Brunswick NJ USA	E1028-500MLGL	to dilute palmitoleic acid
Odorant Cartridge Micromanipulator	Siskiyou, Grants Pass OR USA	MX130R	to position the olfactometer
Flow Vision software	Alicat, Tuscon AZ USA	FLOWVISIONSC	software to control flow rate
Mass Controller	Alicat, Tuscon AZ USA	MC-2SLPM-D	to control the flow rate for humidified air
Mass Controller	Alicat, Tuscon AZ USA	MC-500SCCM-D	to control the flow rate for odor stimulation
Clampex	Molecular Devices, Sunnyvale CA USA	Ver. 10.4	Data acquisition software
Air delivery tube	Ace Glass, Vineland NJ USA	8802-936	to deliver humidified air
50X objective lens	Olympus, Shinjuku Tokyo Japan	LMPLFL50X	recording rig
Clampfit 10	Molecular Devices, Sunnyvale CA USA	Ver. 10.4	software for spike analysis
Igor Pro 6	WaveMetrics, Lake Oswego OR USA	Ver. 6.37	software for data analysis
Audio Monitor	ALA Scientific Instruments, Farmingdale NY USA	NPIEXB-AUDIS-08B	Aurally reports individual spikes
Extracellular Amplifier	ALA Scientific Instruments, Farmingdale NY USA	NPIEXT-02F	to increase the amplitude of electrical signals
Valve Controller	Warner Instruments	VC-8	to control the opening of the valve for odor stimulation
Recording Electrode Micromanipulator	Sutter Instruments, Novato CA USA	MP-285	to position recording electrode
Headstage Amplifier	ALA Scientific Instruments, Farmingdale NY USA	EQ-16.0008	to increase the amplitude of electrical signals
Oscilloscope	Tektronix, Beaverton OR USA	TDS2000C	Visual report of individual spikes