A single cell clonal line which responds reversibly to nerve growth factor (NGF) has been established from a transplantable rat adrenal pheochromocytoma. This line, designated PC12, has a homogeneous and near-diploid chromosome number of 40. By 1 week's exposure to NGF, PC12 cells cease to multiply and begin to extend branching varicose processes similar to those produced by sympathetic neurons in primary cell culture. By several weeks of exposure to NGF, the PC12 processes reach 500-1000 mum in length. Removal of NGF is followed by degeneration of processes within 24 hr and by resumption of cell multiplication within 72 hr. PC12 cells grown with or without NGF contain dense core chromaffin-like granules up to 350 nm in diameter. The NGF-treated cells also contain small vesicles which accumulate in process varicosities and endings. PC12 cells synthesize and store the catecholamine neurotransmitters dopamine and norepinephrine. The levels (per mg of protein) of catecholamines and of the their synthetic enzymes in PC12 cells are comparable to or higher than those found in rat adrenals. NGF-treatment of PC12 cells results in no change in the levels of catecholamines or of their synthetic enzymes when expressed on a per cell basis, but does result in a 4- to 6-fold decrease in levels when expressed on a per mg of protein basis. PC12 cells do not synthesize epinephrine and cannot be induced to do so by treatment with dexamethasone. The PC12 cell line should be a useful model system for neurobiological and neurochemical studies.

https://www.pnas.org/content/pnas/73/7/2424.full.pdf

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

The programmed cell death that occurs as part of normal mammalian development was first observed nearly a century ago (Collin 1906). It has since been established that approximately half of all neurons in the neuroaxis and >99.9% of the total number of cells generated during the course of a human lifetime go on to die through a process of apoptosis (for review, see Datta and Greenberg 1998; Vaux and Korsmeyer 1999). The induction of developmental cell death is a highly regulated process and can be suppressed by a variety of extracellular stimuli. The purification in the 1950s of the nerve growth factor (NGF), which promotes the survival of sympathetic neurons, set the stage for the discovery that peptide trophic factors promote the survival of a wide variety of cell types in vitro and in vivo (Levi-Montalcini 1987). The profound biological consequences of growth factor (GF) suppression of apoptosis are exemplified by the critical role of target-derived neurotrophins in the survival of neurons and the maintenance of functional neuronal circuits. (Pettmann and Henderson 1998). Recently, the ability of trophic factors to promote survival have been attributed, at least in part, to the phosphatidylinositide 38-OH kinase (PI3K)/c-Akt kinase cascade. Several targets of the PI3K/c-Akt signaling pathway have been recently identified that may underlie the ability of this regulatory cascade to promote survival. These substrates include two components of the intrinsic cell death machinery, BAD and caspase 9, transcription factors of the forkhead family, and a kinase, IKK, that regulates the NF-kB transcription factor. This article reviews the mechanisms by which survival factors regulate the PI3K/c-Akt cascade, the evidence that activation of the PI3K/c-Akt pathway promotes cell survival, and the current spectrum of c-Akt targets and their roles in mediating c-Akt-dependent cell survival.

/pdf/cellular-survival-a-play-in-three-akts-3wtxbzcnm7.pdf

Cellular survival: a play in three Akts

Neurotrophins regulate development, maintenance, and function of vertebrate nervous systems. Neurotrophins activate two different classes of receptors, the Trk family of receptor tyrosine kinases and p75NTR, a member of the TNF receptor superfamily. Through these, neurotrophins activate many signaling pathways, including those mediated by ras and members of the cdc-42/ras/rho G protein families, and the MAP kinase, PI-3 kinase, and Jun kinase cascades. During development, limiting amounts of neurotrophins function as survival factors to ensure a match between the number of surviving neurons and the requirement for appropriate target innervation. They also regulate cell fate decisions, axon growth, dendrite pruning, the patterning of innervation and the expression of proteins crucial for normal neuronal function, such as neurotransmitters and ion channels. These proteins also regulate many aspects of neural function. In the mature nervous system, they control synaptic function and synaptic plasticity, while continuing to modulate neuronal survival.

Neurotrophins: roles in neuronal development and function.

Cell Death During Development of the Nervous System

"Embryogenesis is in some way a model system. It has always been distinguished by the exactitude even punctitio, of its anatomical descriptions. An experiment by one of the great masters of embryology could be made the text of a discourse on scientific method. But something is wrong, or has been wrong. There is no theory of development in the sense in which Mendelism is a theory that accounts for the results of breeding experiments. There has therefore been little sense of progression or timeliness about embryological research. Of many papers delivered at embryological meetings, however good they may be in themselves . . . one too often feels that they might have been delivered five years beforehand without making anyone much the wiser, or deferred for five years without making anyone conscious of a great loss" (1). This feeling of frustration so incisively conveyed by these considerations by P. Medawar, pervaded, in the forties, the field of experimental embryology which had been enthusiastically acclaimed in the mid-thirties, when the upper lip of the amphibian blastopore brought this area of research to the forefront of the biological stage. The side branch of experimental neuroembryology, which had stemmed out from the common tree and was entirely devoted to the study of the tropic interrelations between neuronal cell populations and between these and the innervated organs and tissues, was then in its initial vigorous growth phase. It in turn suffered from a sharp decrease in the enthusiasm that had inflamed the pioneers in this field, ever since R. G. Harrison delivered his celebrated lecture on this topic at the Royal Society in London in 1935 (2). Although the alternate "wax and wane" cycles are the rule rather than the exception in all fields of human endeavor, in that of biological sciences the "wane" is all too often indicative of a justified loss of faith in the rational and methodical approach that had at first raised so much hope. A brief account of the state-of-the-art of experimental neuroembryology in the

https://science.sciencemag.org/content/sci/237/4819/1154.full.pdf

The nerve growth factor 35 years later

Nerve growth factor (NGF) is a signaling protein [4] and growth factor implicated in a wide range of development and maintenance functions. NGF was discovered through a series of experiments in the 1950s on the development of the chick [5] nervous system. Since its discovery, NGF has been found to act in a variety of tissues throughout development and adulthood. It has been implicated in immune function, stress response, nerve maintenance, and in neurodegenerative diseases. It is named for its effect on the critical role it plays in the growth and organization [6] of the nervous system during embryonic development.

The nerve-growth factor.

In this paper Viktor Hamburger [5] and Rita Levi-Montalcini [6] collaborated to examine the effects of limb transplantation [7] and explantation on neural development [8]. In 1947 Hamburger invited Levi-Montalcini to his lab at Washington University [9] in St. Louis to examine this question. Independently, each had previously arrived at opposing conclusions based on the same data. Hamburger concluded that limb transplantations caused the ganglia to develop more connections and grow larger while LeviMontalcini concluded that the ganglia first produce a large amount of neurons, then degenerate the unsuccessful neurons. She concluded that larger ganglia must be due to the increase in successful connections. This joint paper, published in the Journal of Experimental Zoology [10] in 1949, corroborated the findings reported by Levi-Montalcini and established that nerve degeneration is an integral part of development.

Proliferation, differentiation and degeneration in the spinal ganglia of the chick embryo under normal and experimental conditions.

In “Selective Growth Stimulating Effects of Mouse Sarcoma on the Sensory and Sympathetic Nervous System of the Chick Embryo,” Rita Levi-Montalcini [5] and Viktor Hamburger [6] explored the effects of two nerve growth stimulating tumors; mouse [7] sarcomas 180 and 37. This experiment led to the discovery that nerve growth factor [8] was a diffusible chemical and later to discoveries that the compound was a protein. Although this paper was an important step in the discovery of nerve growth factor [8], the term “nerve growth factor” was not used in this paper. It was instead referred to as a “growth promoting agent.” The discovery of nerve growth factor [8] earned Levi-Montalcini and Stanley Cohen [9], who also discovered epidermal growth factor, the 1986 Nobel Prize in Physiology or Medicine [10].

Selective growth stimulating effects of mouse sarcoma on the sensory and sympathetic nervous system of the chick embryo.

"Embryogenesis is in some way a model system. It has always been distinguished by the exactitude even puctilio, of its anatomical descriptions. An experiment by one of the great masters of embryology could be made the text of a discourse on scientific method. But something is wrong, or has been wrong. There is no theory of development in the sense in which Mendelism is a theory that accounts for the results of breeding experiments. There has therefore been little sense of progression or timeliness about embryological research. Of many papers delivered at embryological meetings, however good they may be in themselves.. .one too often feels that they might have been delivered five years beforehand without making anyone much the wiser, or deferred for five years without making anyone conscious of a great loss". (1) This feeling of frustration so incisively conveyed by these considerations by P. Medawar, prevaded, in the forties, the field of experimental embryology which had been enthusiastically acclaimed in the mid-thirties, when the upper lip of the amphibian blastopore brought this area of research to the forefront of the biological stage. The side branch of experimental neuroembryology, which had stemmed out from the common tree and was entirely devoted to the study of the trophic interrelations between neuronal cell populations and between these and the innervated organs and tissues was then in its initial vigorous growth phase. It in turn suffered from a sharp decrease in the enthusiasm that had inflamed the pioneers in this field, ever since R. G. Harrison delivered his celebrated lecture on this topic at the Royal Society in London in 1935 (2). Although the alternate 'wax and wane' cycles are the rule rather than the exception in all fields of human endeavor, in that of biological sciences the 'wane' is all too often indicative of a justified loss of faith in the rational and methodical approach that had at first raised so much hope. A brief account of the state-of-the-art of experimental neuroembryology in the forties, when interest in this approach to the study of the developing nervous system was waning, is a prerequisite for understanding the sudden unforseeable turn of events which resulted in the discovery of the Nerve Growth Factor. EXPERIMENTAL NEUROEMBRYOLOGY IN THE EARLY FORTIES

/pdf/the-nerve-growth-factor-thirty-five-years-later-1xj6rh45ve.pdf

Rita Levi-Montalcini

Papers

The nerve growth factor 35 years later

The nerve-growth factor.

Proliferation, differentiation and degeneration in the spinal ganglia of the chick embryo under normal and experimental conditions.

Selective growth stimulating effects of mouse sarcoma on the sensory and sympathetic nervous system of the chick embryo.

The nerve growth factor: thirty-five years later.