Release, storage and uptake of catecholamines by a clonal cell line of nerve growth factor (NGF) responsive pheochromocytoma cells

doi:10.1016/0006-8993(77)90005-1

Brain Research

Volume 129, Issue 2, 1 July 1977, Pages 247-263

https://doi.org/10.1016/0006-8993(77)90005-1 Get rights and content

Abstract

Release, storage and uptake of catecholamines have been studied in cultures of a clonal line of rat pheochromocytoma cells designated as PC12. The PC12 line has previously been shown to respond to nerve growth factor (NGF) by extending neuronal-like processes and to synthesize and contain norepinephrine (NE) and dopamine (DA). In the present experiments, upon exposure to51.5mMK⁺, PC12 cells released a substantial proportion (ca. 30% and 12% for NGF-untreated and -treated cells, respectively) of their endogenous NE and DA. The release was dependent on the presence of Ca²⁺ and was inhibited by excess Mg²⁺. Veratridine, an alkaloid which activates Na⁺ action potential ionophores, was also effective in releasing DA and NE. Exposure of PC12 cells (NGF-treated and -untreated) to reserpine (10⁻⁵ M) resulted in depletion of intracellular DA and NE levels by over 90% in 21 h. PC12 cells were also found to take up NE from the external medium by means of a saturable transport mechanism which follows Michaelis-Menten kinetics (apparentK_m≈ 2 μM), is energy and sodium dependent and which is blocked by low concentrations of cocaine and desmethylimipramine. Such findings indicate that PC12 cells (a) can release catecholamines by means of an exocytotic secretion mechanism, (b) store most of their endogenous catecholamines in chromaffin granules and/or vesicles and (c) have an uptake₁-type transport system for norepinephrine. PC12 cells thus appear to express a number of differentiated properties characteristic of sympathetic neurons and adrenal chromaffin cells and may be a useful system in which to study catecholamine metabolism.

Reference (44)

BreakefieldX.O.
Neurotransmitter metabolism in murine neuroblastoma cells
Life Sci.
(1976)
EvansM.H.
Tetrodotoxin, saxitoxin, and related substances: their applications in neurobiology
Int. Rev. Neurobiol.
(1972)
GoldsteinM. et al.
Changes in catecholamine synthesizing enzyme activities during neuronal growth and degeneration
LowryO.H. et al.
Protein measurement with the Folin phenol reagent
J. biol. Chem.
(1951)
MandelP. et al.
Neuroblastoma cells as a tool for neuronal molecular biology
SangiahS. et al.
Actions of GABA, picrotoxin, and bicuculline on adrenal medulla
Europ. J. Pharmacol.
(1974)
AlmgrenO. et al.
Recovery of dopamine in peripheral adrenergic nerves after reserpine treatment
Naunyn-Schmiedeberg's Arch. exp. Path. Pharmak.
(1976)
AnagnosteB. et al.
Dopamine-β-hydroxylase activity in mouse neuroblastoma tumors and in cell cultures
AntonA.H. et al.
A study of the factors affecting the aluminum oxide-trihydroxyindole procedure for the analysis of catecholamines
J. Pharmacol. exp. Ther.
(1962)
BocchiniV. et al.
The nerve growth factor: Purification as a 30, 000-molecular-weight protein

BornsteinM.D.

Reconstituted rat-tail collagen used as substrate for tissue cultures on coverslips in Maximow slides and roller tubes

Lab. Invest.

(1958)

BoullinD.J.

The action of extracellular cations on the release of the sympathetic transmitter from peripheral nerves

J. Physiol. (Lond.)

(1967)

BreakefieldX.O.

Reserpine sensitivity of catecholamine metabolism in murine neuroblastoma clone NIE-115

J. Neurochem.

(1975)

CatterallW.A. et al.

Sodium uptake associated with activation of action potential ionophores of cultured neuroblastoma and muscle cells

ChalfieM. et al.

Studies of a transplantable rat pheochromocytoma: biochemical characterization and catecholamine secretion

J. Pharmacol. exp. Ther.

(1976)

DouglasW.W.

Stimulus-secretion coupling: the concept and clues from chromaffin and other cells

Brit. J. Pharmacol.

(1968)

DouglasW.W. et al.

The mechanism of catecholamine release from the adrenal medulla and the role of calcium stimulus-secretion coupling

J. Physiol. (Lond.)

(1963)

DouglasW.W. et al.

Stimulant action of barium on the adrenal medulla

Nature (Lond.)

(1964)

GreeneL.A.

A dissociated cell culture bioassay for nerve growth factor

Neurobiology

(1974)

GreeneL.A. et al.

Neuronal properties of hybrid neuroblastoma x sympathetic ganglion cells

GreeneL.A. et al.

Establishment of a noradrenergic clonal line of rat adrenal pheochromocytoma cells which respond to nerve growth factor

HamprechtB.

Neuron models

Angew. Chem.

(1976)

Cited by (346)

Insight into mitochondrial dysfunction mediated by clozapine-induced inhibition of PGRMC1 in PC12 cells
2023, Toxicology
Clozapine is usually considered as the last resort for treatment-resistant schizophrenia (TRS). However, it shows limited efficacy in cognition improvement. Moreover, the metabolic side effects induced by clozapine can aggravate cognitive impairment, which is closely related to its neurotoxicity. Nevertheless, the mechanisms underlying clozapine’s neurotoxicity remain largely elusive. In this study, PC12 cells were simultaneously treated with different concentrations (0 μM, 10 μM, 20 μM, 40 μM and 80 μM) of clozapine and AG205 which functions as a blocking reagent of progesterone receptor membrane component 1 (PGRMC1). In addition, we examined the effect of PGRMC1 in clozapine-induced neurotoxicity through overexpressing or downregulating PGRMC1. Molecular docking and surface plasmon resonance (SPR) analysis indicated that clozapine and AG205 inhibited the binding of endogenous progesterone to PGRMC1. The results showed that high concentration of clozapine and AG205 induced a significant increase in cytotoxicity, reactive oxygen species (ROS) accumulation and mitochondrial membrane potential (MMP) collapse, all of which were worsened as concentration increases, while overexpression of PGRMC1 reverted the above toxic effect of clozapine on PC12 cells. Furthermore, clozapine and AG205 also downregulated the expression of PGRMC1, glucagon-like peptide-1 receptor (GLP-1R) and mitofusin2 (Mfn2). Interestingly, overexpression of PGRMC1 could revert these effects. Our data suggest that overexpression of PGRMC1 in PC12 cells prevents and restores clozapine-induced oxidative and mitochondrial damage. We propose PGRMC1 activation as a promising therapeutic strategy for clozapine-induced neurotoxicity to facilitate the relief of neuronal damage.
Ultra-sensitive surface enhanced Raman spectroscopy sensor for in-situ monitoring of dopamine release using zipper-like ortho-nanodimers
2021, Biosensors and Bioelectronics
Accurate quantitative detection of dopamine (DA) in blood is essential for the early diagnosis and the pathogenesis analysis of dopaminergic dysfunction, which still remains a great challenge because of the extremely low concentration in patients. Using our previously reported DNA-assisted synthesis of ortho-nanodimers (DaSON) strategy, a microfluidic surface enhanced Raman spectroscopy (SERS) biosensor for the ultrasensitive and reliable detection of DA in serum was demonstrated by modifying SERS probes with DA aptamers in a specific orientation to form zipper-like ortho-nanodimers. The uniform 1-nm gap in zipper-like ortho-nanodimers endows the SERS sensor with ultrahigh sensitivity and high accuracy for the detection of DA. The limit of detection is as low as 10 aM in phosphate buffer saline and 10 fM in serum, which is about two orders of magnitude lower than that of previous methods. Using a single microfluidic chip containing a 3D cell culture unit, quantitatively in-situ monitoring of extracellular DA released from living neurons under different medications was first realized. Quantification of DA in human blood samples was also achieved with the recoveries ranging from 87.5% to 123.7%. Given the difficulty of DA quantification in complex physiological samples, our developed SERS sensor provides an appealing tool for in-vitro investigating of neurological processes and clinical examination of dopaminergic disorders.
Cysteine string proteins
2020, Progress in Neurobiology
Citation Excerpt :
Of course, beyond the species difference (cow versus rat), a major factor is likely to be the difference in secretory granule dynamics between these cell types. Whereas chromaffin cells maintain a large and relatively stable pool of granules (see discussion in Graham and Burgoyne, 2000), PC12 cells tonically secrete catecholamines (Greene and Rein, 1977a, b). This is important, because it means that newly synthesized CSP constructs have a higher probability of being incorporated into newly formed secretory organelles in the PC12 cells than in the chromaffin cells.
Cysteine string protein (CSP) was discovered by use of a synapse-specific, monoclonal antibody to screen a cDNA expression library in Drosophila. A vertebrate CSP homolog was later identified and shown to co-purify with synaptic vesicles. CSP-α is now recognized as a membrane constituent of many regulated secretory organelles. Knockout of the csp gene in Drosophila produced temperature-sensitive paralysis reflecting a loss of evoked (but not spontaneous) transmitter release. However, CSP’s role in regulated exocytosis remains ambiguous. Fruit flies lacking the csp gene also exhibited nerve terminal degeneration as did mice deficient in the csp-α gene. This phenotype has been ascribed to the depletion of a functional pool of the t-SNARE, SNAP-25. However, recent studies showing that an endosomal pool of CSP-α contributes to a novel, protein-export pathway argues that CSP’s role in neurodegeneration is more complex. Clients of this later pathway include tau and α-synuclein, proteins linked to neurodegeneration. Additionally, mutations in the csp-α gene cause an adult-onset, neuronal ceroid lipofuscinosis and diminished CSP-α expression is an early event in Alzheimer’s disease. Collectively, these findings indicate that much remains to be learned about the role of CSPs in cellular secretory pathways and human disease.
Model systems for analysis of dopamine transporter function and regulation
2019, Neurochemistry International
The dopamine transporter (DAT) plays a critical role in dopamine (DA) homeostasis by clearing transmitter from the extraneuronal space after vesicular release. DAT serves as a site of action for a variety of addictive and therapeutic reuptake inhibitors, and transport dysfunction is associated with transmitter imbalances in disorders such as schizophrenia, attention deficit hyperactive disorder, bipolar disorder, and Parkinson disease. In this review, we describe some of the model systems that have been used for in vitro analyses of DAT structure, function and regulation, and discuss a potential relationship between transporter kinetic values and membrane cholesterol.
Cellular models as tools for the study of the role of alpha-synuclein in Parkinson's disease
2017, Experimental Neurology
Neurodegenerative diseases are highly debilitating conditions characterised primarily by progressive neuronal loss and impairment of the nervous system. Parkinson's disease (PD) is one of the most common of these disorders, affecting 1–2% of the population above the age of 65. Although the underlying mechanisms of PD have been extensively studied, we still lack a full understanding of the molecular underpinnings of the disease. Thus, the in vitro and in vivo models currently used are able to only partially recapitulate the typical phenotypes of the disease. Here, we review various cell culture models currently used to study the molecular basis of PD, with a focus on alpha-synuclein-associated molecular pathologies. We also discuss how different cell models may constitute powerful tools for high-throughput screening of molecules capable of modulating alpha-synuclein toxicity.
Polychlorinated biphenyls, polybrominated biphenyls, polychlorinated dibenzo-p-dioxins, and polychlorinated dibenzofurans
2017, Reproductive and Developmental Toxicology
Polychlorinated biphenyls (PCBs), polybrominated biphenyls, polychlorinated dibenzo-p-dioxins, and polychlorinated dibenzofurans belong to a group of compounds that are structurally related and are environmentally and biologically persistent. These chemicals are ubiquitous and detectable in a variety of environmental media and biota. Exposure to these chemicals has been linked to a broad spectrum of effects. Fetal and early developmental exposures are particularly sensitive and can have different outcomes when compared to exposure in adults. Latent effects of early exposures include, but are not limited to, depressed circulating thyroid hormone levels and abnormal thyroid cytology; developmental effects of the heart, palate, and kidney; delayed cognitive development; altered sensory and motor abilities; and reproductive impairment and compromised neuronal function. Although AhR activation has been attributed to several dioxin-like coplanar compounds, some PCBs that are noncoplanar in nature seem to exert their toxic effects through different mechanisms, including calcium signaling, oxidative stress, thyroid hormone perturbations, and neurotransmitter imbalance. The most significant problem caused by these compounds involve accidental poisoning via food supply or consumption of contaminated food from contaminated areas. Additionally, there are areas of the environment that are heavily contaminated by these chemicals because of past industrial activities. Animals and humans residing in or near contaminated locations certainly are at risk of serious health effects. Efforts must continue to reduce exposure to protect wildlife and humans. The best way to accomplish is to modernize technological processes to prevent the release of these chemicals into the environment.

View all citing articles on Scopus

View full text

Release, storage and uptake of catecholamines by a clonal cell line of nerve growth factor (NGF) responsive pheochromocytoma cells

Abstract

Life Sci.

Int. Rev. Neurobiol.

J. biol. Chem.

Europ. J. Pharmacol.

Recovery of dopamine in peripheral adrenergic nerves after reserpine treatment

Naunyn-Schmiedeberg's Arch. exp. Path. Pharmak.

Dopamine-β-hydroxylase activity in mouse neuroblastoma tumors and in cell cultures

A study of the factors affecting the aluminum oxide-trihydroxyindole procedure for the analysis of catecholamines

J. Pharmacol. exp. Ther.

The nerve growth factor: Purification as a 30, 000-molecular-weight protein

Reconstituted rat-tail collagen used as substrate for tissue cultures on coverslips in Maximow slides and roller tubes

Lab. Invest.

The action of extracellular cations on the release of the sympathetic transmitter from peripheral nerves

J. Physiol. (Lond.)

Reserpine sensitivity of catecholamine metabolism in murine neuroblastoma clone NIE-115

J. Neurochem.

Sodium uptake associated with activation of action potential ionophores of cultured neuroblastoma and muscle cells

Studies of a transplantable rat pheochromocytoma: biochemical characterization and catecholamine secretion

J. Pharmacol. exp. Ther.

Stimulus-secretion coupling: the concept and clues from chromaffin and other cells

Brit. J. Pharmacol.

The mechanism of catecholamine release from the adrenal medulla and the role of calcium stimulus-secretion coupling

J. Physiol. (Lond.)

Stimulant action of barium on the adrenal medulla

Nature (Lond.)

A dissociated cell culture bioassay for nerve growth factor

Neurobiology

Neuronal properties of hybrid neuroblastoma x sympathetic ganglion cells

Establishment of a noradrenergic clonal line of rat adrenal pheochromocytoma cells which respond to nerve growth factor

Neuron models

Angew. Chem.