Measurements of intracellular ionized calcium in squid giant axons using calcium-selective electrodes

DiPolo, Reinaldo; Rojas, Héctor; Vergara, Julio L.; López, Rubén; Caputo, Carlo

doi:10.1016/0005-2736(83)90500-x

Cited by 36 publications

(18 citation statements)

References 20 publications

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“…However, it should be noted that the [Ca2+]i value measured with Ca electrodes represents only that in a restricted region near the electrode tip which is presumably placed at a substantial distance from the active membrane. Since it is known that the Ca > concentration or Ca 2 § buffering capacity is not uniform within the cytosol (Baker et al, 1971;Rose & Loewenstein, 1975;Loewenstein, 1976;Tillotson & Gorman, 1980;DiPolo et al, 1983;Gorman et al, 1984), the actual Ca dependency of the Ca2+-activated K + channel at the active membrane would be somewhat different from that shown in Fig. 5.…”

Section: Discussionmentioning

confidence: 85%

Oscillations of cytoplasmic concentrations of Ca2+ and K+ in fused L cells

1986

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Using Ca2+- and K+-selective microelectrodes, the cytosolic free Ca2+ and K+ concentrations were measured in mouse fibroblastic L cells. When the extracellular Ca2+ concentration exceeded several micromoles, spontaneous oscillations of the intracellular free Ca2+ concentration were observed in the submicromolar ranges. During the Ca2+ oscillations, the membrane potential was found to oscillate concomitantly. The peak of cyclic increases in the free Ca2+ level coincided in time with the peak of periodic hyperpolarizations. Both oscillations were abolished by reducing the extracellular Ca2+ concentration down to 10(-7) M or by applying a Ca2+ channel blocker, nifedipine (50 microM). In the presence of 0.5 mM quinine, an inhibitor of Ca2+-activated K+ channel, sizable Ca2+ oscillations still persisted, while the potential oscillations were markedly suppressed. Oscillations of the intracellular K+ concentration between about 145 and 140 mM were often associated with the potential oscillations. The minimum phase of the K+ concentration was always 5 to 6 sec behind the peak hyperpolarization. Thus, it is concluded that the oscillation of membrane potential results from oscillatory increases in the intracellular Ca2+ level, which, in turn, periodically stimulate Ca2+-activated K+ channels.

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Section: Discussionmentioning

confidence: 85%

Oscillations of cytoplasmic concentrations of Ca2+ and K+ in fused L cells

1986

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“…Cyanide blocks evoked synaptic transmission in 20-30 min and this rate of block is dependent on presynaptic nerve activity. However, in the axon, substantial recovery of the intracellular calcium concentration from cyanide poisoning usually occurs within 5 min of wash-out (Baker et al 1971;DiPolo et al 1983). The slow recovery of evoked transmission at the synapse may reflect the time required for the nerve terminal to recover from high levels of free calcium.…”

Section: Discussionmentioning

confidence: 99%

“…Lithium sea water effectively depresses calcium buffering in squid axoplasm (Baker & Schlaepfer, 1978; P. F. Baker & J. Umbach, unpublished observation) and can increase (beyond 1 ,UM) the ionized calcium at the periphery of intact axons (Baker, Blaustein, Hodgkin & Steinhardt, 1969;Baker et al 1971;DiPolo et al 1983). On return to normal (sodium) sea water, the high level of ionized calcium is reduced to resting values within minutes.…”

Section: Discussionmentioning

confidence: 99%

“…In the squid axon, cyanide poisoning begins to raise ionized calcium within 20-40 min of application (Baker et al 1971) and the calcium concentration eventually equilibrates at or above the micromolar range (DiPolo, Rojas, Vergara, Lopez & Caputo, 1983;Baker & Umbach, 1983). The rate of rise and final level of ionized calcium can be enhanced by previous loading of calcium into the nerve and by elevating extracellular calcium during poisoning.…”

Section: Discussionmentioning

confidence: 99%

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Inhibitors of calcium buffering depress evoked transmitter release at the squid giant synapse.

Adams¹,

Takeda²,

Umbach³

1985

The Journal of Physiology

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SUMMARY1. Evoked release of transmitter at the squid giant synapse was examined under conditions where the calcium ion concentration in the presynaptic terminal was manipulated by inhibitors of calcium sequestration.2. Simultaneous intracellular recordings of presynaptic and post-synaptic resting and action potentials were made during bath application of one of the following metabolic inhibitors: sodium cyanide (NaCN), carbonyl cyanide-p-trifluoromethoxyphenyl hydrazone (FCCP); ruthenium red (RuR) and sodium-free (lithium) sea water.3. Cyanide and lithium sea water reversibly depressed the post-synaptic potential (p.s.p.) whilst RuR and FCCP blocked the evoked post-synaptic response irreversibly. The progressive reduction of p.s.p. amplitude was accompanied by a reversible increase in synaptic delay.4. The time course of block of the p.s.p. was similar for different agents and dependent on the rate of presynaptic activity (30-40 min at 0-01 Hz). Recovery of the post-synaptic action potential following block by cyanide and lithium sea water was obtained within 40 min and 5 min respectively. 5. Synaptic depression by the metabolic inhibitors does not result from changes in presynaptic resting or action potentials, nor from a change in post-synaptic receptor sensitivity. The post-synaptic response to the local ionophoresis of L-glutamate was unchanged following inhibition of evoked release of transmitter by cyanide. 6. Injections of EGTA into presynaptic terminals poisoned by cyanide produced transient increases in p.s.p. amplitude, suggesting that cyanide is having its effect through raising intracellular calcium rather than lowering ATP. Control experiments injecting EGTA into unpoisoned nerve terminals showed no apparent effect on evoked transmitter release.

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“…The free [Ca 2+ ] was measured with custom-made Ca 2+ electrodes [2,6,32] previously calibrated with pCa (-log 10 [Ca 2+ ]) standard solutions (Calbuff-1, WPI, New Haven, Conn., USA). Only those electrodes that showed linear behavior in the pCa range of from 2 to 8 were used.…”

Section: Equilibrium Properties Of Dm-nmentioning

confidence: 99%

Kinetic properties of DM-nitrophen and calcium indicators: rapid transient response to flash photolysis

Escobar

Vélez

Kim

et al. 1997

Pfl�gers Archiv European Journal of Physiology

130

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We describe a high temporal resolution confocal spot microfluorimetry setup which makes possible the detection of fluorescence transients elicited by Ca2+ indicators in response to large (50-200 microM), short duration (< 100 ns), free [Ca2+] transients generated by laser flash photolysis of DM-nitrophen (DM-n; caged Ca2+). The equilibrium and kinetic properties of the commercially available indicators Fluo-3, Rhod-2, CalciumOrange-5N (COr-5N) and CalciumGreen-2 (CGr-2) were determined experimentally. The data reveal that COr-5N displays simple, fast response kinetics while, in contrast, Fluo-3, Rhod-2 and CGr-2 are characterized by significantly slower kinetic properties. These latter indicators may be unsuitable for tracking Ca2+ signaling events lasting only a few milliseconds. A model which accurately predicts the time course of fluorescence transients in response to rapid free [Ca2+] changes was developed. Experimental data and model predictions concur only when the association rate constant of DM-n is approximately 20 times faster than previously reported. This work establishes a quantitative theoretical framework for the study of fast Ca2+ signaling events and the use of flash photolysis in cells and model systems.

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Measurements of intracellular ionized calcium in squid giant axons using calcium-selective electrodes

Cited by 36 publications

References 20 publications

Oscillations of cytoplasmic concentrations of Ca2+ and K+ in fused L cells

Oscillations of cytoplasmic concentrations of Ca2+ and K+ in fused L cells

Inhibitors of calcium buffering depress evoked transmitter release at the squid giant synapse.

Kinetic properties of DM-nitrophen and calcium indicators: rapid transient response to flash photolysis

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