New Approaches for Analysis of Amperometrical Recordings

Gómez, José Luís Graña; Brioso, Miguel A.; Machado, José David; Sánchez, José Luis; Borges, Ricardo

doi:10.1111/j.1749-6632.2002.tb04544.x

Cited by 23 publications

(18 citation statements)

References 12 publications

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“…Amperometric records were analyzed on a single spike basis in IGOR Pro with a modified peak detection routine based on the "Spike" macro initially described by the Borges group (Gomez et al, 2002 …”

Section: Discussionmentioning

confidence: 99%

Cortical F-Actin, the Exocytic Mode, and Neuropeptide Release in Mouse Chromaffin Cells Is Regulated by Myristoylated Alanine-rich C-Kinase Substrate and Myosin II

Doreian

Fulop

Meklemburg

et al. 2009

MBoC

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Adrenal medullary chromaffin cells are innervated by the sympathetic splanchnic nerve and translate graded sympathetic firing into a differential hormonal exocytosis. Basal sympathetic firing elicits a transient kiss-and-run mode of exocytosis and modest catecholamine release, whereas elevated firing under the sympathetic stress response results in full granule collapse to release catecholamine and peptide transmitters into the circulation. Previous studies have shown that rearrangement of the cell actin cortex regulates the mode of exocytosis. An intact cortex favors kiss-and-run exocytosis, whereas disrupting the cortex favors the full granule collapse mode. Here, we investigate the specific roles of two actin-associated proteins, myosin II and myristoylated alanine-rich C-kinase substrate (MARCKS) in this process. Our data demonstrate that MARCKS phosphorylation under elevated cell firing is required for cortical actin disruption but is not sufficient to elicit peptide transmitter exocytosis. Our data also demonstrate that myosin II is phospho-activated under high stimulation conditions. Inhibiting myosin II activity prevented disruption of the actin cortex, full granule collapse, and peptide transmitter release. These results suggest that phosphorylation of both MARCKS and myosin II lead to disruption of the actin cortex. However, myosin II, but not MARCKS, is required for the activity-dependent exocytosis of the peptide transmitters.

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Section: Discussionmentioning

confidence: 99%

Cortical F-Actin, the Exocytic Mode, and Neuropeptide Release in Mouse Chromaffin Cells Is Regulated by Myristoylated Alanine-rich C-Kinase Substrate and Myosin II

Doreian

Fulop

Meklemburg

et al. 2009

MBoC

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show abstract

“…To ensure synchronization with capacitance measurements, the same signal was also acquired simultaneously in the Pulse software as a second channel. Amperometric records were analyzed with a modified peak detection routine based on the Spike macro (Gomez et al, 2002). We modified the original macro to include analysis of "spike feet."…”

Section: Methodsmentioning

confidence: 99%

Activity-Dependent Differential Transmitter Release in Mouse Adrenal Chromaffin Cells

Fulop¹,

Radabaugh²,

Smith³

2005

J. Neurosci.

202

284

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Chromaffin cells of the adrenal medulla are a primary neuroendocrine output of the sympathetic nervous system. When stimulated, they secrete a host of transmitter molecules, including catecholamines and neuropeptides, through the fusion of dense core secretory granules with the cell surface. At basal firing rates, set by the sympathetic tone, chromaffin cells selectively release catecholamines at a modest rate. Stress-mediated sympathetic activation leads to elevated catecholamine secretion and also evokes neuropeptide release. Catecholamines and neuropeptides are copackaged in the same granules; thus, it is unclear how this activity-dependent differential transmitter release is achieved. In this report, we use electrophysiological, electrochemical, fluorescence, and immunocytochemical approaches to quantify transmitter release under physiological electrical stimulation at the single cell level. We provide data to show that chromaffin cells selectively release catecholamine under basal firing conditions but release both neuropeptides and catecholamines under conditions that match acute stress. We further show that this differential transmitter release is achieved through a regulated activity-dependent dilation of the granule fusion pore. Thus, chromaffin cells may regulate release of different transmitters through a simple size-exclusion mechanism.

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“…Image and data analysis of fluorescence images were performed using custom-written macros in IGOR Pro (WaveMetrics). Single amperometric spikes were analyzed in IGOR Pro with a macro modified from the original "Spike" package (Gomez et al, 2002 …”

Section: Methodsmentioning

confidence: 99%

Myosin II Activation and Actin Reorganization Regulate the Mode of Quantal Exocytosis in Mouse Adrenal Chromaffin Cells

Doreian¹,

Fulop²,

Smith³

2008

J. Neurosci.

108

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Chromaffin cells of the adrenal medulla are innervated by the sympathetic nervous system. Stimulation causes chromaffin cells to fire action potentials, leading to the exocytosis of various classes of transmitters into the circulation. Low-frequency electrical stimulation (action potentials delivered at 0.5 Hz) causes adrenal chromaffin cells to selectively release catecholamines through a kiss-and-run fusion event. Elevated electrical stimulation (action potentials at 15 Hz) evokes fusion pore dilation, full granule collapse, and additional release of the neuropeptide-containing proteinaceous granule core. Here we apply single-cell electrophysiological, electrochemical, and fluorescence measurements to investigate the cellular mechanism for this shift in exocytic behavior. We show that at low-frequency stimulation, a filamentous-actin cell cortex plays a key role in stabilizing the kiss-and-run fusion event. Increased stimulation disrupts the actin cortex, driving full granule collapse. We show that pharmacological perturbation of the actin cortex supersedes stimulus frequency in controlling exocytic mode. Finally, we show that nonmuscle myosin II activation contributes to the cytoskeleton-dependent control of the fusion event. Inhibition of myosin II or myosin light chain kinase under elevated stimulation frequencies inhibits fusion pore dilation and maintains the granule in a kiss-and-run mode of exocytosis. These results demonstrate an essential role for activity-evoked cytoskeletal rearrangement and the action of myosin II in the regulation of catecholamine and neuropeptide exocytosis and represent an essential element of the sympathetic stress response.

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New Approaches for Analysis of Amperometrical Recordings

Cited by 23 publications

References 12 publications

Cortical F-Actin, the Exocytic Mode, and Neuropeptide Release in Mouse Chromaffin Cells Is Regulated by Myristoylated Alanine-rich C-Kinase Substrate and Myosin II

Cortical F-Actin, the Exocytic Mode, and Neuropeptide Release in Mouse Chromaffin Cells Is Regulated by Myristoylated Alanine-rich C-Kinase Substrate and Myosin II

Activity-Dependent Differential Transmitter Release in Mouse Adrenal Chromaffin Cells

Myosin II Activation and Actin Reorganization Regulate the Mode of Quantal Exocytosis in Mouse Adrenal Chromaffin Cells

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