AM-loading of fluorescent Ca2+ indicators into intact single fibers of frog muscle.
AuthorsZhao M, Hollingworth S, Baylor SM
JournalBiophys J
PubMed ID9168048
'The AM loading of a number of different fluorescent Ca2+ indicators was compared in intact single fibers of frog muscle. Among the 13 indicators studied, loading rates (the average increase in the fiber concentration of indicator per first 60 min of loading) varied approximately 100-fold, from approximately 3 microM/h to ... More
Significance of Na/Ca exchange for Ca2+ buffering and electrical activity in mouse pancreatic beta-cells.
AuthorsGall D, Gromada J, Susa I, Rorsman P, Herchuelz A, Bokvist K
JournalBiophys J
PubMed ID10096898
'We have combined the patch-clamp technique with microfluorimetry of the cytoplasmic Ca2+ concentration ([Ca2+]i) to characterize Na/Ca exchange in mouse beta-cells and to determine its importance for [Ca2+]i buffering and shaping of glucose-induced electrical activity. The exchanger contributes to Ca2+ removal at [Ca2+]i above 1 microM, where it accounts for ... More
Photolysis-induced suppression of inhibition in rat hippocampal CA1 pyramidal neurons.
AuthorsWang J, Zucker RS
JournalJ Physiol
PubMed ID11410632
'1. Whole cell patch clamp recording, Ca(2+) measurement with ratiometric fluorescent dyes and photolysis of caged Ca(2+) were combined to investigate the depolarization- and photolysis-induced suppression of inhibition (DSI and PSI) in rat hippocampal CA1 pyramidal cells. 2. A 5-s depolarization from -70 mV to 0 mV or a 6-s ... More
'Cytosolic calcium ([Ca2+]i) is an important mediator of neuronal signal transduction, participating in diverse biochemical reactions that elicit changes in synaptic efficacy, metabolic rate, and gene transcription. Excessive [Ca2+]i also has been implicated as a cause of acute neuronal injury, although measurement of [Ca2+]i in living neurons by fluorescent calcium ... More
Ionic selectivity of low-affinity ratiometric calcium indicators: mag-Fura-2, Fura-2FF and BTC.
AuthorsHyrc KL, Bownik JM, Goldberg MP
JournalCell Calcium
PubMed ID10756974
'Accurate measurement of elevated intracellular calcium levels requires indicators with low calcium affinity and high selectivity. We examined fluorescence spectral properties and ionic specificity of three low-affinity, ratiometric indicators structurally related to Fura-2: mag-Fura-2 (furaptra), Fura-2FF, and BTC. The indicators differed in respect to their excitation wavelengths, affinity for Ca2+ ... More
Confocal Ca2+ imaging of organelles, cells, tissues, and organs.
AuthorsWilliams DA, Bowser DN, Petrou S
JournalMethods Enzymol
PubMed ID10506988
The development of a new family of intracellular calcium probes.
AuthorsIatridou H, Foukaraki E, Kuhn MA, Marcus EM, Haugland RP, Katerinopoulos HE
JournalCell Calcium
PubMed ID8149419
Four new potential fluorescent probes for Ca2+ using the coumarin moiety as a fluorophore have been synthesized and their spectral properties and binding affinities for Ca2+ determined. The most promising derivative for intracellular use, BTC, exhibits an excitation wavelength shift from 462 nm to 401 nm on binding Ca2+, with ... More
Fluorescence-based Tl(+)-influx assays as a novel approach for characterization of small-conductance Ca(2+)-activated K (+) channel modulators.
AuthorsJørgensen S, Johansen TH, Dyhring T,
JournalMethods Mol Biol
PubMed ID18998099
Small-conductance Ca(2+)-activated potassium (SK) channels constitute a family of ion channels that are regulated by the cytosolic Ca(2+) concentration. Increases in the intracellular Ca(2+) concentration ([Ca(2+)](i)) result in opening of the channels, which in turn will lead to changes in the membrane potential. As the name implies, the channels are ... More
A novel assay of Gi/o-linked G protein-coupled receptor coupling to potassium channels provides new insights into the pharmacology of the group III metabotropic glutamate receptors.
AuthorsNiswender CM, Johnson KA, Luo Q, Ayala JE, Kim C, Conn PJ, Weaver CD,
JournalMol Pharmacol
PubMed ID18171729
The group III metabotropic glutamate receptors (mGluRs) represent a family of presynaptically expressed G-protein-coupled receptors (GPCRs) with enormous therapeutic potential; however, robust cellular assays to study their function have been difficult to develop. We present here a new assay, compatible with traditional high-throughput screening platforms, to detect activity of pharmacological ... More
A thallium-sensitive, fluorescence-based assay for detecting and characterizing potassium channel modulators in mammalian cells.
Potassium channels have been identified as targets for a large number of therapeutic indications. The ability to use a high-throughput functional assay for the detection and characterization of small-molecule modulators of potassium channels is very desirable. However, present techniques capable of screening very large chemical libraries are limited in terms ... More
Simultaneous dual-excitation ratiometry using orthogonal linear polarized lights.
AuthorsFukano T, Shimozono S, Miyawaki A
JournalBiochem Biophys Res Commun
PubMed ID15047150
Dual-excitation ratiometric dyes permit quantitative Ca2+ measurements by minimizing the effects of several artifacts that are unrelated to changes in the concentration of free Ca2+ ([Ca2+]). These dyes are excited alternately at two different wavelengths, and the pair of intensity measurements must be collected sequentially. Therefore, it is difficult to ... More
Micromolar and submicromolar Ca2+ spikes regulating distinct cellular functions in pancreatic acinar cells.
AuthorsIto K, Miyashita Y, Kasai H
JournalEMBO J
PubMed ID9029145
Agonists induce Ca2+ spikes, waves and oscillations initiating at a trigger zone in exocrine acinar cells via Ca2+ release from intracellular Ca2+ stores. Using a low affinity ratiometric Ca2+ indicator dye, benzothiazole coumarin (BTC), we found that high concentrations of agonists transiently increased Ca2+ concentrations to the micromolar range (>10 ... More
Kinetic control of multiple forms of Ca(2+) spikes by inositol trisphosphate in pancreatic acinar cells.
AuthorsIto K, Miyashita Y, Kasai H
JournalJ Cell Biol
PubMed ID10427093
The mechanisms of agonist-induced Ca(2+) spikes have been investigated using a caged inositol 1,4,5-trisphosphate (IP(3)) and a low-affinity Ca(2+) indicator, BTC, in pancreatic acinar cells. Rapid photolysis of caged IP(3) was able to reproduce acetylcholine (ACh)-induced three forms of Ca(2+) spikes: local Ca(2+) spikes and submicromolar (<1 microM) and micromolar ... More
Neuronal free calcium measurement using BTC/AM, a low affinity calcium indicator.
AuthorsHyrc KL, Bownik JM, Goldberg MP
JournalCell Calcium
PubMed ID9883271
BTC is a low affinity calcium indicator (Kd approximately 7-26 microM) featuring many desirable properties for cellular calcium imaging, including long excitation wavelengths (400/485 nm), low sensitivity to Mg2+, and accuracy of ratiometric measurement [Iatridou H., Foukaraki E., Kuhn M.A., Marcus E.M., Haugland R.P., Katerinopoulos H.E. The development of a ... More
Presynaptic target of Ca2+ action on neuropeptide and acetylcholine release in Aplysia californica.
1. When buccal neuron B2 of Aplysia californica is co-cultured with sensory neurons (SNs), slow peptidergic synapses are formed. When B2 is co-cultured with neurons B3 or B6, fast cholinergic synapses are formed. 2. Patch pipettes were used to voltage clamp pre- and postsynaptic neurons and to load the caged ... More
Calcium transients in cerebellar granule cell presynaptic terminals.
AuthorsRegehr WG, Atluri PP
JournalBiophys J
PubMed ID7612860
Calcium ions act presynaptically to modulate synaptic strength and to trigger neurotransmitter release. Here we detect stimulus-evoked changes in residual free calcium ([Ca2+]i) in rat cerebellar granule cell presynaptic terminals. Granule cell axons, known as parallel fibers, and their associated boutons, were labeled with several calcium indicators. When parallel fibers ... More
Development of a Novel Lead that Targets M. tuberculosis Polyketide Synthase 13.
Authors
JournalCell
PubMed ID28669536
Spirocycle MmpL3 Inhibitors with Improved hERG and Cytotoxicity Profiles as Inhibitors of Mycobacterium tuberculosis Growth.