Biomedical subjects

Morris Feldman

Publications and source records attributed to Morris Feldman.

4 recordsLinked to original sources

PubMed2006

Ca2+ -induced Ca2+ release through localized Ca2+ uncaging in smooth muscle.

Ca(2+)-induced Ca(2+) release (CICR) from the sarcoplasmic reticulum (SR) occurs in smooth muscle as spontaneous SR Ca(2+) release or Ca(2+) sparks and, in some spiking tissues, as Ca(2+) release that is triggered by the activation of sarcolemmal Ca(2+) channels. Both processes display spatial localization in that release occurs at a higher frequency at specific subcellular regions. We have used two-photon flash photolysis (TPFP) of caged Ca(2+) (DMNP-EDTA) in Fluo-4-loaded urinary bladder smooth muscle cells to determine the extent to which spatially localized increases in Ca(2+) activate SR release and to further understand the molecular and biophysical processes underlying CICR. TPFP resulted in localized Ca(2+) release in the form of Ca(2+) sparks and Ca(2+) waves that were distinguishable from increases in Ca(2+) associated with Ca(2+) uncaging, unequivocally demonstrating that Ca(2+) release occurs subsequent to a localized rise in [Ca(2+)](i). TPFP-triggered Ca(2+) release was not constrained to a few discharge regions but could be activated at all areas of the cell, with release usually occurring at or within several microns of the site of photolysis. As expected, the process of CICR was dominated by ryanodine receptor (RYR) activity, as ryanodine abolished individual Ca(2+) sparks and evoked release with different threshold and kinetics in FKBP12.6-null cells. However, TPFP CICR was not completely inhibited by ryanodine; Ca(2+) release with distinct kinetic features occurred with a higher TPFP threshold in the presence of ryanodine. This high threshold release was blocked by xestospongin C, and the pharmacological sensitivity and kinetics were consistent with CICR release at high local [Ca(2+)](i) through inositol trisphosphate (InsP(3)) receptors (InsP(3)Rs). We conclude that CICR activated by localized Ca(2+) release bears essential similarities to those observed by the activation of I(Ca) (i.e., major dependence on the type 2 RYR), that the release is not spatially constrained to a few specific subcellular regions, and that Ca(2+) release through InsP(3)R can occur at high local [Ca(2+)](i).

Animals↗

PubMed2005

Gene trap and gene inversion methods for conditional gene inactivation in the mouse.

Conditional inactivation of individual genes in mice using site-specific recombinases is an extremely powerful method for determining the complex roles of mammalian genes in developmental and tissue-specific contexts, a major goal of post-genomic research. However, the process of generating mice with recombinase recognition sequences placed at specific locations within a gene, while maintaining a functional allele, is time consuming, expensive and technically challenging. We describe a system that combines gene trap and site-specific DNA inversion to generate mouse embryonic stem (ES) cell clones for the rapid production of conditional knockout mice, and the use of this system in an initial gene trap screen. Gene trapping should allow the selection of thousands of ES cell clones with defined insertions that can be used to generate conditional knockout mice, thereby providing extensive parallelism that eliminates the time-consuming steps of targeting vector construction and homologous recombination for each gene.

Animals↗

PubMed2004

Methodologic advancements in the study of airway smooth muscle.

The study of isolated airway myocytes has provided important information relative to specific processes that regulate contraction, proliferation, and synthetic properties of airway smooth muscle (ASM). To place this information in physiological context, however, improved methods to examine airway biology in vivo are needed. Advances in genetic, biochemical, and optical methods provide unprecedented opportunities to improve our understanding of in vivo physiology and pathophysiology. This article describes 4 important methodologic advances in the study of ASM: (1) the development of transgenic mice that could be used to investigate ASM proliferation and phenotype switching during the development of hypersensitivity, and to investigate excitation-contraction coupling; (2) the use of CD38-deficient mice to confirm the role of CD38-dependent, cyclic adenosine diphosphate-ribose-mediated calcium release in airway responsiveness; (3) investigation of the role of actin filament length and p38 mitogen-activated protein kinase activity in regulating the mechanical plasticity-elasticity balance in contracted ASM; and (d) the use of bronchial biopsies to study ASM structure and phenotype in respiratory science.

ADP-ribosyl Cyclase↗

PubMed2002

PECAM-1 (CD31) regulates a hydrogen peroxide-activated nonselective cation channel in endothelial cells.

Hydrogen peroxide (H2O2) released by neutrophils is an important mediator of endothelial cell (EC) injury and vascular inflammation via its effect on EC-free Ca2+, [Ca2+]i. Although the underlying mechanisms are not well understood, platelet endothelial cell adhesion molecule (PECAM)-1/CD-31 is a critical modulator of neutrophil-EC transmigration. PECAM-1 is also known to regulate EC calcium signals and to undergo selective tyrosine phosphorylation. Here, we report that PECAM-1 molecules transduce EC responses to hydrogen peroxide. In human umbilical vein EC and REN cells (a PECAM-1-negative EC-like cell line) stably transfected with PECAM-1 (RHP), noncytolytic H2O2 exposure (100-200 microM H2O2) activated a calcium-permeant, nonselective cation current, and a transient rise in [Ca2+]i of similar time course. Neither response was observed in untransfected REN cells, and H2O2-evoked cation current was ablated in REN cells transfected with PECAM-1 constructs mutated in the cytoplasmic tyrosine-containing domain. The PECAM-dependent H2O2 current was inhibited by dialysis of anti-PECAM-1 cytoplasmic domain antibodies, required Src family tyrosine kinase activity, was independent of inositol trisphosphate receptor activation, and required only an intact PECAM-1 cytoplasmic domain. PECAM-1-dependent H2O2 currents and associated [Ca2+]i transients may play a significant role in regulating neutrophil-endothelial interaction, as well as in oxidant-mediated endothelial response and injury.

Calcium↗