Gelsolin displaces phalloidin from actin filaments. A new fluorescence method shows that both Ca2+ and Mg2+ affect the rate at which gelsolin severs F-actin.
AuthorsAllen PG, Janmey PA
JournalJ Biol Chem
PubMed ID7806519
'We describe an assay for measuring both the extent and kinetics of the severing of F-actin, based on the enhanced fluorescence emission of tetramethylrhodamine-phalloidin bound to F-actin. The enhanced fluorescence is lost after exposure to active gelsolin by displacement of the phalloidin from actin during severing. This assay requires small ... More
Optical measurement of presynaptic calcium currents.
AuthorsSabatini BL, Regehr WG
JournalBiophys J
PubMed ID9512051
'Measurements of presynaptic calcium currents are vital to understanding the control of transmitter release. However, most presynaptic boutons in the vertebrate central nervous system are too small to allow electrical recordings of presynaptic calcium currents (I(Ca)pre). We therefore tested the possibility of measuring I(Ca)pre optically in boutons loaded with calcium-sensitive ... More
Measurement of intracellular free zinc in living cortical neurons: routes of entry.
'We used the ratioable fluorescent dye mag-fura-5 to measure intracellular free Zn2+ ([Zn2+]i) in cultured neocortical neurons exposed to neurotoxic concentrations of Zn2+ in concert with depolarization or glutamate receptor activation and identified four routes of Zn2+ entry. Neurons exposed to extracellular Zn2+ plus high K+ responded with a peak ... More
Presynaptic strontium dynamics and synaptic transmission.
AuthorsXu-Friedman MA, Regehr WG
JournalBiophys J
PubMed ID10096899
'Strontium can replace calcium in triggering neurotransmitter release, although peak release is reduced and the duration of release is prolonged. Strontium has therefore become useful in probing release, but its mechanism of action is not well understood. Here we study the action of strontium at the granule cell to Purkinje ... More
Fluorescent zinc indicators for neurobiology.
AuthorsThompson RB, Peterson D, Mahoney W, Cramer M, Maliwal BP, Suh SW, Frederickson C, Fierke C, Herman P
JournalJ Neurosci Methods
PubMed ID12191759
'Mounting evidence indicates that zinc has multiple roles in cell biology, viz. as a part of metalloenzyme catalytic sites, as a structural component of gene regulatory proteins, and (like calcium) as a free signal ion, particularly in the cortex of the brain. While most Zn(II) in the brain is tightly ... More
The regulation of intracellular Mg2+ in guinea-pig heart, studied with Mg(2+)-selective microelectrodes and fluorochromes.
AuthorsBuri A, Chen S, Fry CH, Illner H, Kickenweiz E, McGuigan JA, Noble D, Powell T, Twist VW
JournalExp Physiol
PubMed ID8471242
'Because of the reported presence of a Na(+)-Mg2+ exchanger in guinea-pig but not in ferret myocardium, the Mg2+ extrusion mechanism in guinea-pig myocardium has been reinvestigated using Mg(2+)- and Na(+)- selective microelectrodes and the fluorochromes mag-fura-2 and -5. The mean [Mg2+]i measured with microelectrodes in trabeculae or papillary muscles was ... More
Binding of phosphate, aluminum fluoride, or beryllium fluoride to F-actin inhibits severing by gelsolin.
AuthorsAllen PG, Laham LE, Way M, Janmey PA
JournalJ Biol Chem
PubMed ID8617730
'Actin exhibits ATPase activity of unknown function that increases when monomers polymerize into filaments. Differences in the kinetics of ATP hydrolysis and the release of the hydrolysis products ADP and inorganic phosphate suggest that phosphate-rich domains exist in newly polymerized filaments. We examined whether the enrichment of phosphate on filamentous ... More
Measurement of intracellular free zinc in living neurons.
AuthorsCanzoniero LM, Sensi SL, Choi DW
JournalNeurobiol Dis
PubMed ID9361304
Excessive Zn2+ influx has been implicated in the pathogenesis of neuronal death after global ischemia or prolonged seizures, but little is presently known about cellular regulation of intracellular free Zn2+ ([Zn2+]i). In large part, this is because the tools currently available for measuring [Zn2+]i are limited in comparison to those ... More
Micromolar Ca2+ transients in dendritic spines of hippocampal pyramidal neurons in brain slice.
AuthorsPetrozzino JJ, Pozzo Miller LD, Connor JA
JournalNeuron
PubMed ID7605633
The magnitude and dynamics of [Ca2+] changes in spines and dendrites of hippocampal CA1 pyramidal neurons have been characterized using a low affinity fluorescent indicator, mag-Fura 5, that is sensitive to Ca2+ in the micromolar range. During tetanic stimulation (1 s), we observed progressive [Ca2+] increases in distal CA1 spines ... More
Evaluation of mag-fura-5, the new fluorescent indicator for free magnesium measurements.
AuthorsIllner H, McGuigan JA, Lüthi D
JournalPflugers Arch
PubMed ID1488274
The new fluorescent indicator, mag-fura-5, was evaluated for its ability to measure accurately physiological changes in cytosolic free magnesium. The apparent dissociation constants (Kd) of the fluorochrome for Mg2+, Mg2+/EGTA and Ca2+/EGTA solutions were 14.7 mM, 15.4 mM, and 1.8 mM respectively. The calculated difference in the fluorescence ratios and ... More
Timing of neurotransmission at fast synapses in the mammalian brain.
AuthorsSabatini BL, Regehr WG
JournalNature
PubMed ID8906792
Understanding the factors controlling synaptic delays has broad implications. On a systems level, the speed of synaptic transmission limits the communication rate between neurons and strongly influences local circuit dynamics. On a molecular level, the delay from presynaptic calcium entry to postsynaptic responses constrains the molecular mechanism of vesicle fusion. ... More
Properties of tri- and tetracarboxylate Ca2+ indicators in frog skeletal muscle fibers.
AuthorsZhao M, Hollingworth S, Baylor SM
JournalBiophys J
PubMed ID8789107
Recently a number of lower-affinity fluorescent Ca2+ indicators have become available with principal absorbance bands at visible wavelengths. This article evaluates these indicators, as well as two shorter wavelength indicators, mag-fura-5 and mag-indo-1, for their suitability as rapid Ca2+ indicators in frog skeletal muscle fibers. With three lower-affinity tricarboxylate indicators ... More
Interplay between sodium and calcium dynamics in granule cell presynaptic terminals.
AuthorsRegehr WG
JournalBiophys J
PubMed ID9370441
Fluorescent indicators were used to detect stimulus-evoked changes in presynaptic levels of intracellular sodium (Na(i)) and calcium (Ca(i)) in granule cell parallel fibers in brain slices from rat cerebellum. Ca(i) increased during stimulation, and three exponentials were needed to approximate its return to prestimulus levels. Ca(i) decayed to approximately 10% ... More
Detecting changes in calcium influx which contribute to synaptic modulation in mammalian brain slice.
AuthorsSabatini BL, Regehr WG
JournalNeuropharmacology
PubMed ID8606793
The control of neurotransmitter release by modulation of presynaptic calcium influx was investigated at the granule cell to Purkinje cell synapse in rat cerebellar slices. Excitatory post-synaptic currents were measured using whole cell voltage clamp, and changes in presynaptic Ca influx were determined with the Ca-sensitive dye mag-fura-5. Single stimuli ... More