Synaptic vesicle cytochemistry changes when cultured sympathetic neurones develop cholinergic interactions

Johnson, Mary I.; Ross, Don; Meyers, Marta; Rees, Rosemary P.; Bunge, Richard P.; Wakshull, Eric; Burton, Harold

doi:10.1038/262308a0

Cited by 128 publications

(62 citation statements)

References 14 publications

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“…The induction of ACh synthesis in cultured sympathetic neurons by heart cell CM is slow, requiring Johnson et al 1976). Given that the same protein can induce the production of several neuropeptides, it was of interest to determine whether the peptide changes followed a time course similar to that of ACh.…”

Section: Time Course Of Peptide Induction and Its Reversibilitymentioning

confidence: 99%

Generation of Neuronal Diversity: Analogies and Homologies with Hematopoiesis

Nawa

Yamamori²,

Le³

et al. 1990

Cold Spring Harbor Symposia on Quantitative Biology

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Section: Time Course Of Peptide Induction and Its Reversibilitymentioning

confidence: 99%

Generation of Neuronal Diversity: Analogies and Homologies with Hematopoiesis

Nawa

Yamamori²,

Le³

et al. 1990

Cold Spring Harbor Symposia on Quantitative Biology

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“…However, if these same neurons are grown in the presence of non-neuronal cells or in medium conditioned by them, the cultures synthesize significant amounts of acetylcholine (AcCh) as well as catecholamines; if sufficiently high concentrations of conditioned medium are used, then the cultures become predominantly cholinergic (2)(3)(4)(5). An interesting question is whether single neurons in these cultures display either exclusively adrenergic or cholinergic properties or whether single neurons possess mixed properties.…”

mentioning

confidence: 99%

Rat sympathetic neurons and cardiac myocytes developing in microcultures: correlation of the fine structure of endings with neurotransmitter function in single neurons.

Landis¹

1976

Proc. Natl. Acad. Sci. U.S.A.

130

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Microcultures containing single sympathetic principal neurons and small numbers of dissociated heart myocytes were prepared from newborn rats. After the transmitter properties of the neuron were studied by electrophysiological experiments, the microculture was examined with the electron microscope. Single neurons of either putative cholinergic or putative adrenergic character made morphological synapses on themselves (autapses), although only cholinergic autapses were detected electrophysiologicaily. Numerous axonal varicosities were present adjacent to the myocytes but no synaptic specializations were evident. After permanganate fixation to localize endogenous norepinephrine, the endings of neurons which appeared to secrete catecholamines contained many small granular vesicles, while the endings of neurons which appeared to secrete acetylcholine contained none. The endings of neurons which apparently secreted both catecholamines and acetylcholine contained only occasional small granular vesicles.If sympathetic neurons, dissociated from the superior cervical ganglia of newborn rats, are grown in the absence of non-neuronal cells, then the cultures are predominantly adrenergic in character (1). However, if these same neurons are grown in the presence of non-neuronal cells or in medium conditioned by them, the cultures synthesize significant amounts of acetylcholine (AcCh) as well as catecholamines; if sufficiently high concentrations of conditioned medium are used, then the cultures become predominantly cholinergic (2-5). An interesting question is whether single neurons in these cultures display either exclusively adrenergic or cholinergic properties or whether single neurons possess mixed properties. In a preliminary series of experiments (6), 3-week-old cultures that synthesized predominantly catecholamines, predominantly AcCh, or significant amounts of both were obtained by using varying proportions of conditioned medium; the incidence of cholinergic synapse formation was estimated electrophysiologically and, varicosities were examined with the electron microscope in sister cultures fixed with permanganate to localize norepinephrine (NE) in small granular vesicles (SGV) (7). In predominantly adrenergic cultures, SGV were absent from only 1% of the endings, but as the amount of AcCh synthesized and the fraction of neurons shown to interact cholinergically increased, the proportion of terminals in the culture lacking SGV also increased. Because some terminals containing numerous SGV and resembling those in predominantly adrenergic cultures were also observed in mixed cultures, it seemed likely that the cultures contained at least two populations of neurons. On the other hand, several observations raised the possibility that some neurons were not exclusively adrenergic or cholinergic: (i) in some cultures, the proportion of terminals which contained no SGV and were presumably cholinergic was less than the proportion of neurons interacting cholinergically; (ii) some terminals with SGV contained relativel...

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“…For example, it is now well established that neurons may alter the transmitters they synthesize and secrete, and thus alter their phenotypes, under appropriate circumstances (5)(6)(7). Alteration of neurotransmitter expression in response to experimental alteration in the environment has been particularly well defined in neural crest-derived sympathetic and sensory neurons.…”

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confidence: 99%

Determination of synaptic phenotype: insulin and cAMP independently initiate development of electrotonic coupling between cultured sympathetic neurons.

Kessler¹,

Spray²,

Sáez³

et al. 1984

Proc. Natl. Acad. Sci. U.S.A.

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Electrotonic coupling between pairs of sympathetic neurons dissociated from superior cervical ganglia of neonatal rats is rare when cells are cultured for 2 weeks in a nutrient medium plus serum and is common when cells are cultured for the same period in serum-free defined medium. This defined medium is the same nutrient medium with five added factors (progesterone, transferrin, putrescine, insulin, and selenium). When added singly to serum-containing medium, insulin and, to a lesser extent, selenium promote the development of electrotonic and dye coupling. The insulin effect is obtained with doses as low as 0.01 jag/nm and is maximal after exposures from 3 to 5 days. The incidence of electrotonic coupling is also enhanced by exposure of cells to dibutyryl cAMP. This effect is obtained with doses as low as 0.1 mM, is faster (being maximal at =12 hr exposure), and is prolonged in the presence of the phosphodiesterase inhibitor caffeine. Butyrate itself promotes coupling to a small extent, but cAMP involvement is confirmed by similar effects of other membrane permeant analogues. Endogenous levels of cAMP are significantly elevated in cultures grown in the defined medium but not in those in serum-containing medium to which insulin or selenium are added. We conclude that the promotion of coupling by cAMP and by insulin or selenium are independent.The development of coupling in the defined medium thus seems to be a consequence of the addition of promoting substances (insulin, selenium) and the removal of an inhibitory effect of serum on cAMP levels.Neuronal differentiation and specialization are influenced by the environment, so that the phenotype of a neuron may depend on the extracellular milieu (1-4). For example, it is now well established that neurons may alter the transmitters they synthesize and secrete, and thus alter their phenotypes, under appropriate circumstances (5-7). Alteration of neurotransmitter expression in response to experimental alteration in the environment has been particularly well defined in neural crest-derived sympathetic and sensory neurons. For example, regions of the neural crest that normally give rise to cholinergic neurons can be induced to develop noradrenergic progeny by transplanting them into a region of the crest from which noradrenergic ganglia normally arise (8). Conversely, sympathetic neurons that in vivo express predominantly noradrenergic characteristics become cholinergic when cultured in the presence of certain non-neuronal cells (5).Phenotypic modifiability is not restricted to choice of transmitter; it also involves other aspects of neuronal differentiation. For example, Higgins and Burton (9) found that embryonic sympathetic neurons cultured in a chemically defined serum-free medium become electrotonically coupled, a mode of synaptic transmission not normally utilized by sympathetic gangliaJ This observation was confirmed by Wolinsky et al. (10). The cultured neurons continue to express noradrenergic characteristics, including tyrosine hydroxylase and dopamin...

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Synaptic vesicle cytochemistry changes when cultured sympathetic neurones develop cholinergic interactions

Cited by 128 publications

References 14 publications

Generation of Neuronal Diversity: Analogies and Homologies with Hematopoiesis

Generation of Neuronal Diversity: Analogies and Homologies with Hematopoiesis

Rat sympathetic neurons and cardiac myocytes developing in microcultures: correlation of the fine structure of endings with neurotransmitter function in single neurons.

Determination of synaptic phenotype: insulin and cAMP independently initiate development of electrotonic coupling between cultured sympathetic neurons.

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