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Morphological control of inositol-1,4,5-trisphosphate-dependent signals.

Fink CC, Slepchenko B, Moraru II, Schaff J, Watras J, Loew LM - J. Cell Biol. (1999)

Bottom Line: We conclude that the characteristic calcium dynamics requires rapid, high-amplitude production of [InsP(3)](cyt) in the neurite.This requisite InsP(3) spatiotemporal profile is provided, in turn, as an intrinsic consequence of the cell's morphology, demonstrating how geometry can locally and dramatically intensify cytosolic signals that originate at the plasma membrane.In addition, the model predicts, and experiments confirm, that stimulation of just the neurite, but not the soma or growth cone, is sufficient to generate a calcium response throughout the cell.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, University of Connecticut Health Center, Farmington, Connecticut 06030, USA.

ABSTRACT
Inositol-1,4,5-trisphosphate (InsP(3))-mediated calcium signals represent an important mechanism for transmitting external stimuli to the cell. However, information about intracellular spatial patterns of InsP(3) itself is not generally available. In particular, it has not been determined how the interplay of InsP(3) generation, diffusion, and degradation within complex cellular geometries can control the patterns of InsP(3) signaling. Here, we explore the spatial and temporal characteristics of [InsP(3)](cyt) during a bradykinin-induced calcium wave in a neuroblastoma cell. This is achieved by using a unique image-based computer modeling system, Virtual Cell, to integrate experimental data on the rates and spatial distributions of the key molecular components of the process. We conclude that the characteristic calcium dynamics requires rapid, high-amplitude production of [InsP(3)](cyt) in the neurite. This requisite InsP(3) spatiotemporal profile is provided, in turn, as an intrinsic consequence of the cell's morphology, demonstrating how geometry can locally and dramatically intensify cytosolic signals that originate at the plasma membrane. In addition, the model predicts, and experiments confirm, that stimulation of just the neurite, but not the soma or growth cone, is sufficient to generate a calcium response throughout the cell.

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Localized BK application. a, Simulations. [Ca2+]cyt is shown for simulations of focal 500 nM BK application to the soma, neurite, and growth cone. b, Experiments. Cells were loaded with fura-2, 340/380 ratio pairs collected with a cooled CCD camera, and results displayed in pseudocolored [Ca2+]. 500-nM BK was focally applied (arrowheads) by pressure ejection from a micropipette in a flow chamber at time = 0 s. Bars, 25 μm. Note that the pseudocolor [Ca2+] scales are not identical.
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Figure 4: Localized BK application. a, Simulations. [Ca2+]cyt is shown for simulations of focal 500 nM BK application to the soma, neurite, and growth cone. b, Experiments. Cells were loaded with fura-2, 340/380 ratio pairs collected with a cooled CCD camera, and results displayed in pseudocolored [Ca2+]. 500-nM BK was focally applied (arrowheads) by pressure ejection from a micropipette in a flow chamber at time = 0 s. Bars, 25 μm. Note that the pseudocolor [Ca2+] scales are not identical.

Mentions: To further probe whether this amplified [InsP3] in the neurite is necessary and sufficient for initiation and propagation of a calcium wave, we modeled the condition of a local BK stimulation in three distinct cell regions: soma, middle of the neurite, and growth cone (or distal neurite). The results of these simulations are shown in Fig. 4 a. When BK is added to only the soma, elevation of calcium levels occur after a long delay, no wave is generated, and peak [Ca2+]cyt is low (∼500 nM). When BK is applied to the neurite, a calcium wave propagates in both directions; whereas [Ca2+]cyt in the neurite is comparable to that produced by global BK application, [Ca2+]cyt in the soma is lower (∼500 nM). Finally, when the application of BK is simulated for only the distal neurite, an immediate calcium increase is seen in that region. However, the wave fails to propagate far up the neurite, and [Ca2+]cyt soon returns to baseline levels. To check these predictions, we performed a series of experiments in which BK was focally applied by pressure ejection to the cells (Fig. 4 b). When BK was focally applied to the soma, a gradual increase of calcium was seen in the soma, which failed to propagate down the neurite as a wave (observed in 7/8 cells). When BK is focally applied to only the neurite, a calcium wave is typically initiated (9/13 cells), although [Ca2+]cyt in the soma didn't reach the levels seen with global BK application. Finally, when BK is focally applied to only the most distal neurite (or growth cone), a local elevation of calcium was observed (7/11 cells), which failed to propagate as a calcium wave to the soma. Together, these experiments validate the predictions made by the simulations, and show that the morphologically enhanced InsP3 signal in the neurite is necessary and sufficient for initiation and propagation of a calcium wave.


Morphological control of inositol-1,4,5-trisphosphate-dependent signals.

Fink CC, Slepchenko B, Moraru II, Schaff J, Watras J, Loew LM - J. Cell Biol. (1999)

Localized BK application. a, Simulations. [Ca2+]cyt is shown for simulations of focal 500 nM BK application to the soma, neurite, and growth cone. b, Experiments. Cells were loaded with fura-2, 340/380 ratio pairs collected with a cooled CCD camera, and results displayed in pseudocolored [Ca2+]. 500-nM BK was focally applied (arrowheads) by pressure ejection from a micropipette in a flow chamber at time = 0 s. Bars, 25 μm. Note that the pseudocolor [Ca2+] scales are not identical.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2169350&req=5

Figure 4: Localized BK application. a, Simulations. [Ca2+]cyt is shown for simulations of focal 500 nM BK application to the soma, neurite, and growth cone. b, Experiments. Cells were loaded with fura-2, 340/380 ratio pairs collected with a cooled CCD camera, and results displayed in pseudocolored [Ca2+]. 500-nM BK was focally applied (arrowheads) by pressure ejection from a micropipette in a flow chamber at time = 0 s. Bars, 25 μm. Note that the pseudocolor [Ca2+] scales are not identical.
Mentions: To further probe whether this amplified [InsP3] in the neurite is necessary and sufficient for initiation and propagation of a calcium wave, we modeled the condition of a local BK stimulation in three distinct cell regions: soma, middle of the neurite, and growth cone (or distal neurite). The results of these simulations are shown in Fig. 4 a. When BK is added to only the soma, elevation of calcium levels occur after a long delay, no wave is generated, and peak [Ca2+]cyt is low (∼500 nM). When BK is applied to the neurite, a calcium wave propagates in both directions; whereas [Ca2+]cyt in the neurite is comparable to that produced by global BK application, [Ca2+]cyt in the soma is lower (∼500 nM). Finally, when the application of BK is simulated for only the distal neurite, an immediate calcium increase is seen in that region. However, the wave fails to propagate far up the neurite, and [Ca2+]cyt soon returns to baseline levels. To check these predictions, we performed a series of experiments in which BK was focally applied by pressure ejection to the cells (Fig. 4 b). When BK was focally applied to the soma, a gradual increase of calcium was seen in the soma, which failed to propagate down the neurite as a wave (observed in 7/8 cells). When BK is focally applied to only the neurite, a calcium wave is typically initiated (9/13 cells), although [Ca2+]cyt in the soma didn't reach the levels seen with global BK application. Finally, when BK is focally applied to only the most distal neurite (or growth cone), a local elevation of calcium was observed (7/11 cells), which failed to propagate as a calcium wave to the soma. Together, these experiments validate the predictions made by the simulations, and show that the morphologically enhanced InsP3 signal in the neurite is necessary and sufficient for initiation and propagation of a calcium wave.

Bottom Line: We conclude that the characteristic calcium dynamics requires rapid, high-amplitude production of [InsP(3)](cyt) in the neurite.This requisite InsP(3) spatiotemporal profile is provided, in turn, as an intrinsic consequence of the cell's morphology, demonstrating how geometry can locally and dramatically intensify cytosolic signals that originate at the plasma membrane.In addition, the model predicts, and experiments confirm, that stimulation of just the neurite, but not the soma or growth cone, is sufficient to generate a calcium response throughout the cell.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology, University of Connecticut Health Center, Farmington, Connecticut 06030, USA.

ABSTRACT
Inositol-1,4,5-trisphosphate (InsP(3))-mediated calcium signals represent an important mechanism for transmitting external stimuli to the cell. However, information about intracellular spatial patterns of InsP(3) itself is not generally available. In particular, it has not been determined how the interplay of InsP(3) generation, diffusion, and degradation within complex cellular geometries can control the patterns of InsP(3) signaling. Here, we explore the spatial and temporal characteristics of [InsP(3)](cyt) during a bradykinin-induced calcium wave in a neuroblastoma cell. This is achieved by using a unique image-based computer modeling system, Virtual Cell, to integrate experimental data on the rates and spatial distributions of the key molecular components of the process. We conclude that the characteristic calcium dynamics requires rapid, high-amplitude production of [InsP(3)](cyt) in the neurite. This requisite InsP(3) spatiotemporal profile is provided, in turn, as an intrinsic consequence of the cell's morphology, demonstrating how geometry can locally and dramatically intensify cytosolic signals that originate at the plasma membrane. In addition, the model predicts, and experiments confirm, that stimulation of just the neurite, but not the soma or growth cone, is sufficient to generate a calcium response throughout the cell.

Show MeSH
Related in: MedlinePlus