Innovative Uses of Axon Microelectrode Amplifiers

See references at the bottom of the page. If you have an innovative use of an Axon Instruments amplifier that you would like to see featured on this page, contact Technical Support.

Axopatch 200B Patch Clamp Amplifier
Measures Presynaptic Calcium Influx - Optically!

Julio Vergara (Department of Physiology, UCLA) and colleagues have taken advantage of the ultra-low noise performance of the cooled-headstage CV-203BU and Axopatch 200B amplifier in patch mode to measure photodiode currents in a scanning confocal spot detection system. They used the calcium indicator Oregon Green 488 BAPTA-5N and argon laser epi-illumination to measure the spatial extent and kinetics of action potential-induced presynaptic calcium domains, which they hypothesize are closely related to the sites of calcium influx responsible for synaptic transmitter release. In addition, the Axopatch 200B is ideal for flash photolysis experiments with this system, as the forced reset input of the amplifier can rapidly (50 µs) reset the capacitive-feedback headstage following the high-intensity UV "uncaging" flash.

Spatial dependence of action potential (AP)-induced presynaptic fluorescence transients. A.  CCD image of a nerve terminal loaded with 600 µM OGB-5N. Right, Camera lucida diagram of the same nerve terminal at higher magnification illustrating the size of the illumination spot (open circle) relative to the nerve terminal. The arrow indicates the direction of the spot displacement, and the asterisk indicates the location in which the largest transient was acquired. B.  A family of single AP-induced OGB-5N fluorescence transients (bottom traces) recorded consecutively at various spot locations in 0.3 µm steps along the horizontal line (double arrow in A). The black trace corresponds to the average of six neuronal APs that elicited fluorescence records. EGTA (50 µM) was included in the pipette solution. Fluorescence traces were filtered at 1 kHz. Two model simulation traces are superimposed on the experimental records (black dotted traces). C.  Three-dimensional plot obtained by offsetting all the fluorescence traces in the scan according to their relative spot-detection location. Color bands are increments of 0.05 DF/F.   Reprinted by permission. Copyright 1999 The Society for Neuroscience.

Low Current Scanning Tunneling Microscopy
with the Axopatch 200B Patch Clamp Amplifier

Ralph Nyffenegger (Park Scientific Instruments, Sunnyvale CA) has interfaced an Axopatch 200B with Park Scientific Instruments Autoprobe CP and VP2 microscopes to enable low-current scanning tunneling microscopy (STM). With the existing current / voltage converter of the instrument, the typical lower limit of the set current was about 50-100 pA. With the addition of the Axopatch 200B, the tunneling current limit was reduced by more than 100-fold, to 0.5 pA. For more information on this technique, see the Focus on Methods article Using the Axopatch 200B for Low-Current Scanning Tunneling Microscopy in AxoBits 23 . The CV-4 headstage has also been used for low-current STM (see Dunlap, et al. reference below).

Hyperbaric Recordings from Rat Brainstem Slices
with the Axoclamp 2A

Jay Dean and Dan Mulkey (Department of Physiology and Biophysics, Wright State University School of Medicine) have used an Axoclamp 2A with HS-2A headstage in a specially constructed recording apparatus to obtain continuous intracellular recordings and pH measurements from rat brainstem slices under changing hyperbaric conditions. These technically difficult recordings make it possible to differentiate effects of pressure per se from those of increased gas partial pressure (e.g., PO2, PN2, PCO2) in the same neuron, and to investigate the reversibility of these effects. Such experiments will provide insight into the cellular mechanisms involved in CNS oxygen toxicity, nitrogen narcosis and carbon dioxide toxicity.

Note: Axon Instruments provides this description of recording from cells at high pressure using the Axoclamp and HS-2 headstage because we think it presents a fascinating application of our instruments. However, we do not endorse the use of our equipment in this way. None of our products are designed for use at high pressure; nor are they tested at high pressure. In particular, we would recommend against using our other headstages at high pressure, because the hybrid integrated circuit inside the headstage would be susceptible to pressure damage.

References

DiGregorio, D.A., Peskoff, A., Vergara, J.L. Measurement of Action Potential-Induced Presynaptic Calcium Domains at a Cultured Neuromuscular Junction. J. Neurosci., 19:7846-7859 (1999).

Escobar, A.L., Velez, P., Kim, A.M., Cifuentes, F., Fill, M., Vergara, J. L. Kinetic properties of DM-nitrophen and calcium indicators: rapid transient response to flash photolysis. Pflügers Arch - Eur J Physiol, 434:615-631 (1997).

Dunlap, D., Smith, S., Bustamante, C., Tamayo, J., García, R. A very low current scanning tunneling microscope. Rev. Sci. Instrum., 66:4876-4879 (1995).

Dean, J.B. and Mulkey, D.K. Continuous intracellular recording from mammalian neurons exposed to hyperbaric helium, oxygen, or air. J. Appl. Physiol., 89:807-822 (2000).

Dean, J.B., Mulkey, D.K., Arehart, J. Details on building a hyperbaric chamber for intracellular recording in brain tissue slices. J. Appl. Physiol., see http://jap.physiology.org/cgi/content/full/89/2/807/DC1 or NAPS Document 05566, National Auxiliary Publications, c/o Microfiche Publications, 248 Hempstead Turnpike, West Hempstead, NY 11552.