Apparent role of traveling metabolic waves in oxidant release by living neutrophils

Abstract

Cell metabolism self-organizes into two types of dissipative structures: chemical oscillations and traveling metabolic waves. In the present study we test the hypothesis that traveling NAD(P)H waves within neutrophils are associated spatially and temporally with the release of reactive oxygen metabolites (ROMs). Using high-speed optical microscopy and taking advantage of the autofluorescence of NAD(P)H, we have observed the propagation of NAD(P)H waves within cells. When NAD(P)H waves reach the lamellipodium of morphologically polarized neutrophils, a diffusing plume of superoxide is released as evidenced by the conversion of hydroethidine in the extracellular environment to ethidium bromide. Parallel results were obtained by using high-speed emission microspectrophotometry. These experiments indicate that the spatial and temporal properties of NAD(P)H waves are transformed into ROM pulses in the extracellular environment. Propagating NAD(P)H waves allow neutrophils to specifically deliver substrate to the lamellipodium at high concentrations, thus facilitating the local and periodic release of ROMs in the direction of cell movement and͞or a target.

Intracellular Calcium Waves Accompany Neutrophil Polarization, Formylmethionylleucylphenylalanine Stimulation, and Phagocytosis: A High Speed Microscopy Study

“…The high speed microscopic imaging techniques outlined in this and previous papers (23)(24)(25)(26) were designed to permit analyses of chemical wave propagation events within immune cells and should be generally applicable. For example, interactions been leukocytes and targets such as microbes and tumor cells is one potential area of interest.…”

“…We speculate that intracellular signaling may be understood in terms of spatiotemporal variables. Because enzymes can respond to Ca 2ϩ , ATP, NADPH, pH, cAMP, and other intracellular conditions and because Ca 2ϩ , NAD(P)H, and pH exhibit various signaling patterns (23)(24)(25)(26), a variety of superimposed spatiotemporal patterns could be generated. Hence, a few chemicals, each with a set of characteristic patterns, could generate many spatiotemporal enzyme activity patterns.…”

“…A Gen-II tube was used to provide maximal efficiency in the violet-blue region of the spectrum (30). The camera was controlled by a high speed Princeton ST-133 interface and a Stanford Research Systems (Sunnyvale, CA) DG-535 delay gate generator (26). To improve computer acquisition times, the size of the pixel array was adjusted.…”

“…We have recently developed a high speed microscopic imaging technique that captures individual images with 50 ns to ms exposure times at up to 1000 frames/s (23)(24)(25)(26), which allows us to collect stop-action movies of extremely rapid cell signaling events.…”

Cutting Edge: Optical Microspectrophotometry Supports the Existence of Gel Phase Lipid Rafts at the Lamellipodium of Neutrophils: Apparent Role in Calcium Signaling

“…To increase light collection efficiency, the microscope's bottom port was fiber-optically coupled to the input side of an Acton-150 (Acton Instruments, Acton, MA) imaging spectrophotometer. The exit side was connected to a liquid N 2 -cooled intensifier attached to a Peltier-cooled I-MAX-512 camera (approximately Ϫ20°C) (Princeton Instruments) (12,13). …”