Isolation and Characterization of Secretory Vesicles from Bovine Neurohypophyses

Gratzl, Manfred; Torp‐Pedersen, Christian; Dartt, Darlene A.; Treiman, Marek; Thorn, Niels A.

doi:10.1515/bchm2.1980.361.2.1615

Cited by 42 publications

(48 citation statements)

References 18 publications

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“…Similar observations have been made in neurohypophysial secretory vesicles (20,24). However, the colocation of cell membrane components within chromaffin vesicles in conjunction with cotransport studies on the intact tissue as well as on chromaffin cells in culture suggest a participation of chromaffin secretory vesicles in the biogenesis of the cell membrane.…”

Section: Fractionsupporting

confidence: 76%

“…During sucrose-gradient centrifugation secretory vesic1es are exposed to hypertonie media, which probably causes the instability of the isolated material. To circumvent this disadvantage secretory vesicles from the neurohypophysis and adrenal medulla have been isolated on isotonic continuous gradients using Percoll as gradient material (6,7,20,21). The main contaminants of cmde secretory vesicle fractions (Le., those obtained by differential centrifugation) are mitochondria, which can be completely removed using such gradients.…”

Section: Resul Ts and Discussionmentioning

confidence: 99%

“…The main contaminants of cmde secretory vesicle fractions (Le., those obtained by differential centrifugation) are mitochondria, which can be completely removed using such gradients. The vesicles obtained are stable and have successfully been used in studies concerned with the analysis of proton, sotium, and calcium transport (6,7,13,20,(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22)(23)(24).…”

Section: Resul Ts and Discussionmentioning

confidence: 99%

“…Reports from severallaboratories have indicated that lysosomal enzyme activities are found also in secretory vesicle fractions after further purification on Percoll gradients (6,20,24).…”

Section: Resul Ts and Discussionmentioning

confidence: 99%

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Distribution of chromaffin secretory vesicles, acetylcholinesterase, and lysosomal enzymes in sucrose and percoll gradients

Gratzl

1984

Analytical Biochemistry

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emde chromaffin secretory vesicles, obtained by differential centrifugation, were further purified on isotonic (PereolI) gradients. The chromaffin vesicIe fractions recovered from the gradients contain acetylcholinesterase as well as lysosomal enzymes. With the aid of a subsequent sucrose gradient lysosomal enzymes could be removed from chromaffin vesicIe fractions. but not acetylcholinesterase. This suggests that lysosomal enzymes do not pass through the chromaffin vesicIes during the biogenesis of lysosomes but acetylcholinesterase does.

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Section: Fractionsupporting

confidence: 76%

Section: Resul Ts and Discussionmentioning

confidence: 99%

Section: Resul Ts and Discussionmentioning

confidence: 99%

“…Reports from severallaboratories have indicated that lysosomal enzyme activities are found also in secretory vesicle fractions after further purification on Percoll gradients (6,20,24).…”

Section: Resul Ts and Discussionmentioning

confidence: 99%

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Distribution of chromaffin secretory vesicles, acetylcholinesterase, and lysosomal enzymes in sucrose and percoll gradients

Gratzl

1984

Analytical Biochemistry

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“…Stellation induced by adenosine, C3 toxin or Y-27632 can be reversed-i.e., spreading can be restored-by subnanomolar concentrations of the neurohormone VP acting at V1a receptors. These findings have physiological relevance because (1) adenosine is readily generated from breakdown of ATP, which is present in VP secretory vesicles (Gratzl et al 1980), (2) exocytosis of these vesicles occurs locally from axon swellings, i.e. in the pituicyte environment (Morris et al 1988), and (3) pituicyte shape in turn is likely to influence neurohormone output (Theodosis and MacVicar 1996;Hatton 1999).…”

Section: Introductionmentioning

confidence: 94%

Pituicyte Stellation is Prevented by RhoA-or Cdc42-Dependent Actin Polymerization

et al. 2007

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Our aim was to shed light on different steps leading from metabotropic receptor activation to changes in cell shape, such as those that characterize the morphological plasticity of neurohypophysial astrocytes (pituicytes). Using explant cultures of adult rat pituicytes, we have previously established that adenosine A1 receptor activation induces stellation via inhibition of RhoA monomeric GTPase and subsequent disruption of actin stress fibers. Here, we rule out RhoA phosphorylation as a mechanism for that inhibition. Rather, our results are more consistent with involvement of a GTPase-activating protein (GAP). siRNA and pull-down experiments suggest that a step downstream of RhoA might involve Cdc42, another GTPase of the Rho family. However, RhoA activation, e.g., in the presence of serum, induces stress fibers, whereas direct Cdc42 activation appears to confine actin within a submembrane-i.e., cortical-network, which also prevents stellation. Therefore, we propose that RhoA may activate Cdc42 in parallel with an effector, such as p160Rho-kinase, that induces and maintains actin stress fibers in a dominant fashion. Racl is not involved in the stellation process per se but appears to induce a dendritogenic effect. Ultimately, it may be stated that pituicyte stellation is inducible upon mere actin depolymerization, and preventable upon actin organization, be it in the form of stress fibers or in a cortical configuration.

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RhoA inhibition is a key step in pituicyte stellation induced by A₁‐type adenosine receptor activation

Rosso¹,

Peteri-Brünback²,

Vouret‐Craviari

et al. 2002

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Pituicyte stellation in vitro represents a useful model with which to study morphological changes that occur in vivo in these cells during times of high neurohypophysial hormone output. This model has helped us establish the hypothesis of a purinergic regulation of pituicyte morphological plasticity. We first show that ATP induces stellation in 37% of pituicytes, an effect that is secondary to the metabolism of ATP to adenosine. Adenosine-induced stellation of pituicytes appears to be mediated by A(1)-type receptors. The effect is independent of intracellular calcium and does not involve the mitogen-activated protein kinase pathway. The basal (nonstellate) state of pituicytes depends on tonic activation of a Rho GTPase because both C3 transferase (a Rho inhibitor) and Y-27632 (an inhibitor of p160Rho kinase) can induce stellation. Lysophosphatidic acid, a Rho activator, blocks the morphogenic effect of adenosine dose-dependently. Using a specific RhoA pull-down assay, we also show that downregulation of activated RhoA is the key event coupling A(1) receptor activation to pituicyte stellation, via F-actin depolymerization and microtubule reorganization. Finally, both vasopressin and oxytocin can prevent or reverse adenosine-induced stellation. The effects of vasopressin, and those of high concentrations of oxytocin, are mediated through V(1a) receptors. Placed within the context of the relevant literature, our data suggest the possibility of a purinergic regulation of pituicyte morphological plasticity and subsequent modulation of hormone release, with these hormones providing a negative feedback mechanism.

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Isolation and Characterization of Secretory Vesicles from Bovine Neurohypophyses

Cited by 42 publications

References 18 publications

Distribution of chromaffin secretory vesicles, acetylcholinesterase, and lysosomal enzymes in sucrose and percoll gradients

Distribution of chromaffin secretory vesicles, acetylcholinesterase, and lysosomal enzymes in sucrose and percoll gradients

Pituicyte Stellation is Prevented by RhoA-or Cdc42-Dependent Actin Polymerization

RhoA inhibition is a key step in pituicyte stellation induced by A₁‐type adenosine receptor activation

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Isolation and Characterization of Secretory Vesicles from Bovine Neurohypophyses

Cited by 42 publications

References 18 publications

Distribution of chromaffin secretory vesicles, acetylcholinesterase, and lysosomal enzymes in sucrose and percoll gradients

Distribution of chromaffin secretory vesicles, acetylcholinesterase, and lysosomal enzymes in sucrose and percoll gradients

Pituicyte Stellation is Prevented by RhoA-or Cdc42-Dependent Actin Polymerization

RhoA inhibition is a key step in pituicyte stellation induced by A1‐type adenosine receptor activation

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RhoA inhibition is a key step in pituicyte stellation induced by A₁‐type adenosine receptor activation