Effects of injecting calcium-buffer solution on [Ca2+]i in voltage-clamped snail neurons

Biophysical Journal
H J Kennedy, R C Thomas

Abstract

We have investigated why fura-2 and Ca(2+)-sensitive microelectrodes report different values for the intracellular free calcium ion concentration ([Ca(2+)]i or its negative log, pCa(i)) of snail neurons voltage-clamped to -50 or -60 mV. Both techniques were initially calibrated in vitro, using calcium calibration solutions that had ionic concentrations similar to those of snail neuron cytoplasm. Pressure injections of the same solutions at resting and elevated [Ca(2+)]i were used to calibrate both methods in vivo. In fura-2-loaded cells these pressure injections generated changes in [Ca(2+)]i that agreed well with those expected from the in vitro calibration. Thus, using fura-2 calibrated in vitro, the average resting [Ca(2+)]i was found to be 38 nM (pCa(i) 7.42 +/- 0.05). With Ca(2+)-sensitive microelectrodes, the first injection of calibration solutions always caused a negative shift in the recorded microelectrode potential, as if the injection lowered [Ca2+]i. No such effects were seen on the fura-2 ratio. When calibrated in vivo the Ca(2+)-sensitive microelectrode gave an average resting [Ca2+]i of approximately 25 nM (pCa(i) 7.6 +/- 0.1), much lower than when calibrated in vitro. We conclude that [Ca(2+)]i in snail neurons...Continue Reading

Citations

Apr 6, 2006·The Journal of Membrane Biology·G KleesP W Dierkes
Aug 27, 2003·The Journal of General Physiology·Johannes ReisertStephan Frings
Feb 12, 1998·The American Journal of Physiology·T L HerringC E Morris
Feb 8, 2005·The Journal of Experimental Biology·Michael S HedrickPhilip E Bickler
Apr 3, 2013·Cold Spring Harbor Protocols·Roger C Thomas, Donald M Bers
Apr 3, 2013·Cold Spring Harbor Protocols·Roger C Thomas, Donald M Bers

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