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Cycling of dense core vesicles involved in somatic exocytosis of serotonin by leech neurons.

Trueta C, Kuffler DP, De-Miguel FF - Front Physiol (2012)

Bottom Line: A partial bleaching of the spots followed by another depolarization in the presence of FM1-43 produced restaining of some spots, other spots disappeared, some remained without restaining and new spots were formed.Several hours after electrical stimulation the FM1-43 spots accumulated at the center of the somata.This correlated with electron micrographs of multivesicular bodies releasing their contents near Golgi apparatuses.

View Article: PubMed Central - PubMed

Affiliation: Instituto Nacional de Psiquiatría "Ramón de la Fuente Muñiz," México D. F., México.

ABSTRACT
We studied the cycling of dense core vesicles producing somatic exocytosis of serotonin. Our experiments were made using electron microscopy and vesicle staining with fluorescent dye FM1-43 in Retzius neurons of the leech, which secrete serotonin from clusters of dense core vesicles in a frequency-dependent manner. Electron micrographs of neurons at rest or after 1 Hz stimulation showed two pools of dense core vesicles. A perinuclear pool near Golgi apparatuses, from which vesicles apparently form, and a peripheral pool with vesicle clusters at a distance from the plasma membrane. By contrast, after 20 Hz electrical stimulation 47% of the vesicle clusters were apposed to the plasma membrane, with some omega exocytosis structures. Dense core and small clear vesicles apparently originating from endocytosis were incorporated in multivesicular bodies. In another series of experiments, neurons were stimulated at 20 Hz while bathed in a solution containing peroxidase. Electron micrographs of these neurons contained gold particles coupled to anti-peroxidase antibodies in dense core vesicles and multivesicular bodies located near the plasma membrane. Cultured neurons depolarized with high potassium in the presence of FM1-43 displayed superficial fluorescent spots, each reflecting a vesicle cluster. A partial bleaching of the spots followed by another depolarization in the presence of FM1-43 produced restaining of some spots, other spots disappeared, some remained without restaining and new spots were formed. Several hours after electrical stimulation the FM1-43 spots accumulated at the center of the somata. This correlated with electron micrographs of multivesicular bodies releasing their contents near Golgi apparatuses. Our results suggest that dense core vesicle cycling related to somatic serotonin release involves two steps: the production of clear vesicles and multivesicular bodies after exocytosis, and the formation of new dense core vesicles in the perinuclear region.

No MeSH data available.


Related in: MedlinePlus

Ultrastructural relationships of perinuclear vesicles with the nucleus (n), Golgi (go), and vesicle clusters (vc). (A) Low magnification image of the soma of a neuron that had been stimulated with 20 Hz trains in a bathing solution in which magnesium was substituted for calcium. Under these conditions, electrical activity fails to evoke somatic exocytosis and vesicle mobilization, and as can be seen, the areas immediate to the plasma membrane (pm) are devoid of vesicle clusters. Scale = 2.0 μm. (B) Golgi apparatus in the perinuclear area surrounded by clear and dense core vesicles. On the right, there are rows of small vesicles (arrows), some of which are clear while others have electron dense material (small arrowheads). More distantly there are small vesicles with dense cores continued by vesicles with large dense cores (large arrowheads). A fragment of a multivesicular body (mvb) can also be seen. On the top left electron dense vesicles align with the layers of the Golgi apparatus (large arrowhead). (C) Higher magnification image of vesicles adjacent to the nucleus, in the vicinity of mitochondria (m) and Golgi. (D) Clear and dense core vesicles are adjacent to the nuclear membrane (nm). A filament projecting from the vesicle toward the nuclear membrane is marked by the arrow and amplified in the inset. Scale bars for (B–D) = 500 nm.
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Figure 2: Ultrastructural relationships of perinuclear vesicles with the nucleus (n), Golgi (go), and vesicle clusters (vc). (A) Low magnification image of the soma of a neuron that had been stimulated with 20 Hz trains in a bathing solution in which magnesium was substituted for calcium. Under these conditions, electrical activity fails to evoke somatic exocytosis and vesicle mobilization, and as can be seen, the areas immediate to the plasma membrane (pm) are devoid of vesicle clusters. Scale = 2.0 μm. (B) Golgi apparatus in the perinuclear area surrounded by clear and dense core vesicles. On the right, there are rows of small vesicles (arrows), some of which are clear while others have electron dense material (small arrowheads). More distantly there are small vesicles with dense cores continued by vesicles with large dense cores (large arrowheads). A fragment of a multivesicular body (mvb) can also be seen. On the top left electron dense vesicles align with the layers of the Golgi apparatus (large arrowhead). (C) Higher magnification image of vesicles adjacent to the nucleus, in the vicinity of mitochondria (m) and Golgi. (D) Clear and dense core vesicles are adjacent to the nuclear membrane (nm). A filament projecting from the vesicle toward the nuclear membrane is marked by the arrow and amplified in the inset. Scale bars for (B–D) = 500 nm.

Mentions: Ultra thin sections from five somata that had been stimulated in the ganglion with trains at 1 Hz, a frequency that does not evoke somatic exocytosis, contained dense core vesicle clusters distributed in two main areas (Figures 1 and 2). A perinuclear region contained vesicles together with Golgi apparatuses, mitochondria, and endoplasmic reticulum. Small clear vesicles were located next to Golgi apparatuses and some small vesicles displayed dense cores. At larger distances from Golgi, vesicles were large and had dense cores. Multivesicular bodies were mainly located adjacent to the perinuclear vesicle clusters and were characterized by their bag shapes and membrane multilayers filled with dense core and clear vesicles (Figure 1, see also below).


Cycling of dense core vesicles involved in somatic exocytosis of serotonin by leech neurons.

Trueta C, Kuffler DP, De-Miguel FF - Front Physiol (2012)

Ultrastructural relationships of perinuclear vesicles with the nucleus (n), Golgi (go), and vesicle clusters (vc). (A) Low magnification image of the soma of a neuron that had been stimulated with 20 Hz trains in a bathing solution in which magnesium was substituted for calcium. Under these conditions, electrical activity fails to evoke somatic exocytosis and vesicle mobilization, and as can be seen, the areas immediate to the plasma membrane (pm) are devoid of vesicle clusters. Scale = 2.0 μm. (B) Golgi apparatus in the perinuclear area surrounded by clear and dense core vesicles. On the right, there are rows of small vesicles (arrows), some of which are clear while others have electron dense material (small arrowheads). More distantly there are small vesicles with dense cores continued by vesicles with large dense cores (large arrowheads). A fragment of a multivesicular body (mvb) can also be seen. On the top left electron dense vesicles align with the layers of the Golgi apparatus (large arrowhead). (C) Higher magnification image of vesicles adjacent to the nucleus, in the vicinity of mitochondria (m) and Golgi. (D) Clear and dense core vesicles are adjacent to the nuclear membrane (nm). A filament projecting from the vesicle toward the nuclear membrane is marked by the arrow and amplified in the inset. Scale bars for (B–D) = 500 nm.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Figure 2: Ultrastructural relationships of perinuclear vesicles with the nucleus (n), Golgi (go), and vesicle clusters (vc). (A) Low magnification image of the soma of a neuron that had been stimulated with 20 Hz trains in a bathing solution in which magnesium was substituted for calcium. Under these conditions, electrical activity fails to evoke somatic exocytosis and vesicle mobilization, and as can be seen, the areas immediate to the plasma membrane (pm) are devoid of vesicle clusters. Scale = 2.0 μm. (B) Golgi apparatus in the perinuclear area surrounded by clear and dense core vesicles. On the right, there are rows of small vesicles (arrows), some of which are clear while others have electron dense material (small arrowheads). More distantly there are small vesicles with dense cores continued by vesicles with large dense cores (large arrowheads). A fragment of a multivesicular body (mvb) can also be seen. On the top left electron dense vesicles align with the layers of the Golgi apparatus (large arrowhead). (C) Higher magnification image of vesicles adjacent to the nucleus, in the vicinity of mitochondria (m) and Golgi. (D) Clear and dense core vesicles are adjacent to the nuclear membrane (nm). A filament projecting from the vesicle toward the nuclear membrane is marked by the arrow and amplified in the inset. Scale bars for (B–D) = 500 nm.
Mentions: Ultra thin sections from five somata that had been stimulated in the ganglion with trains at 1 Hz, a frequency that does not evoke somatic exocytosis, contained dense core vesicle clusters distributed in two main areas (Figures 1 and 2). A perinuclear region contained vesicles together with Golgi apparatuses, mitochondria, and endoplasmic reticulum. Small clear vesicles were located next to Golgi apparatuses and some small vesicles displayed dense cores. At larger distances from Golgi, vesicles were large and had dense cores. Multivesicular bodies were mainly located adjacent to the perinuclear vesicle clusters and were characterized by their bag shapes and membrane multilayers filled with dense core and clear vesicles (Figure 1, see also below).

Bottom Line: A partial bleaching of the spots followed by another depolarization in the presence of FM1-43 produced restaining of some spots, other spots disappeared, some remained without restaining and new spots were formed.Several hours after electrical stimulation the FM1-43 spots accumulated at the center of the somata.This correlated with electron micrographs of multivesicular bodies releasing their contents near Golgi apparatuses.

View Article: PubMed Central - PubMed

Affiliation: Instituto Nacional de Psiquiatría "Ramón de la Fuente Muñiz," México D. F., México.

ABSTRACT
We studied the cycling of dense core vesicles producing somatic exocytosis of serotonin. Our experiments were made using electron microscopy and vesicle staining with fluorescent dye FM1-43 in Retzius neurons of the leech, which secrete serotonin from clusters of dense core vesicles in a frequency-dependent manner. Electron micrographs of neurons at rest or after 1 Hz stimulation showed two pools of dense core vesicles. A perinuclear pool near Golgi apparatuses, from which vesicles apparently form, and a peripheral pool with vesicle clusters at a distance from the plasma membrane. By contrast, after 20 Hz electrical stimulation 47% of the vesicle clusters were apposed to the plasma membrane, with some omega exocytosis structures. Dense core and small clear vesicles apparently originating from endocytosis were incorporated in multivesicular bodies. In another series of experiments, neurons were stimulated at 20 Hz while bathed in a solution containing peroxidase. Electron micrographs of these neurons contained gold particles coupled to anti-peroxidase antibodies in dense core vesicles and multivesicular bodies located near the plasma membrane. Cultured neurons depolarized with high potassium in the presence of FM1-43 displayed superficial fluorescent spots, each reflecting a vesicle cluster. A partial bleaching of the spots followed by another depolarization in the presence of FM1-43 produced restaining of some spots, other spots disappeared, some remained without restaining and new spots were formed. Several hours after electrical stimulation the FM1-43 spots accumulated at the center of the somata. This correlated with electron micrographs of multivesicular bodies releasing their contents near Golgi apparatuses. Our results suggest that dense core vesicle cycling related to somatic serotonin release involves two steps: the production of clear vesicles and multivesicular bodies after exocytosis, and the formation of new dense core vesicles in the perinuclear region.

No MeSH data available.


Related in: MedlinePlus