Innervation of heart cells in culture by an endogenous source of cholinergic neurons.

Lane, Michelle A.; Sastre, Antonio; Salpeter, Miriam M.

doi:10.1073/pnas.73.12.4506

Cited by 10 publications

(4 citation statements)

References 25 publications

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“…Cholinergic neurons were identified previously in cocultures of embryonic mouse heart but only in preparations from hearts collected at 9 days in utero (20). Viable neurons were not detected in cocultures from later embryonic ages or from 1-day postnatal hearts.…”

Section: Discussionmentioning

confidence: 79%

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Phenotypic properties of adult mouse intrinsic cardiac neurons maintained in culture

Hoard

Hoover²,

Wondergem³

2007

American Journal of Physiology-Cell Physiology

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Intrinsic cardiac neurons are core elements of a complex neural network that serves as an important integrative center for regulation of cardiac function. Although mouse models are used frequently in cardiovascular research, very little is known about mouse intrinsic cardiac neurons. Accordingly, we have dissociated neurons from adult mouse heart, maintained these cells in culture, and defined their basic phenotypic properties. Neurons in culture were primarily unipolar, and 89% had prominent neurite outgrowth after 3 days (longest neurite length of 258 +/- 20 microm, n = 140). Many neurites formed close appositions with other neurons and nonneuronal cells. Neurite outgrowth was drastically reduced when neurons were kept in culture with a majority of nonneural cells eliminated. This finding suggests that nonneuronal cells release molecules that support neurite outgrowth. All neurons in coculture showed immunoreactivity for a full complement of cholinergic markers, but about 21% also stained for tyrosine hydroxylase, as observed previously in sections of intrinsic cardiac ganglia from mice and humans. Whole cell patch-clamp recordings demonstrated that these neurons have voltage-activated sodium current that is blocked by tetrodotoxin and that neurons exhibit phasic or accommodating patterns of action potential firing during a depolarizing current pulse. Several neurons exhibited a fast inward current mediated by nicotinic ACh receptors. Collectively, this work shows that neurons from adult mouse heart can be maintained in culture and exhibit appropriate phenotypic properties. Accordingly, these cultures provide a viable model for evaluating the physiology, pharmacology, and trophic factor sensitivity of adult mouse cardiac parasympathetic neurons.

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Section: Discussionmentioning

confidence: 79%

“…Functional criteria were used to establish the cholinergic phenotype of intrinsic cardiac neurons obtained from embryonic hearts (20). These preparations contained clusters of beating myocytes that had visible connections with clusters of neurons.…”

Section: Discussionmentioning

confidence: 99%

Phenotypic properties of adult mouse intrinsic cardiac neurons maintained in culture

Hoard

Hoover²,

Wondergem³

2007

American Journal of Physiology-Cell Physiology

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“…These neurons, which migrate along the vagus nerves (Mitchell, 1953), reach the heart at an early stage in gestation, i.e. 9 days in the mouse (Lane et al, 1976). The neurons associated with the trachea have been studied more recently by histochemistry (Grillo & Nadel, 1980;Baker et al, 1986) and by scanning and transmission electron microscopy (Cameron & Coburn, 1984;BaIuk et al, 1985;Chiang & Gabella, 1986), and it has been shown that these ganglia extend over the bronchi .…”

Section: Neuron Distribution and Connectivitymentioning

confidence: 96%

Fine structure of the autonomic ganglia of the mouse pulmonary vein

Bałuk

Gabella

1987

J Neurocytol

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The aim of this study was to describe the architecture of a ganglionated nerve plexus found in the loose connective tissue surrounding the pulmonary vein of the mouse. The input to this plexus was from the vagus nerves and from the stellate ganglia. A large ganglion containing more than 200 neurons was commonly found near the primary bifurcation of the pulmonary vein. The neurons were studied by NADH-diaphorase, zinc iodide-osmium and glyoxylic acid-induced catecholamine fluorescence methods at the light microscopic level, by scanning electron microscopy after the removal of connective tissue, and by transmission electron microscopy. The shape of the neuronal cell bodies was generally a smooth ellipsoid with the average major axis about twice the minor axis. The measured maximum cell diameter ranged from 14 to 42 micron (mean 26 micron). The profile area of individual neurons, as measured from wholemount preparations, ranged from 100 to 800 micron2 (mean 340 micron2) and the calculated neuronal volume ranged from 500 to 12,000 micron3 (mean 3300 micron3). Although there was this wide spread in neuronal size, histograms of cell size showed no separate populations of neurons. Almost all of the ganglionic neuronal cell bodies showed no catecholamine-specific fluorescence, but about 1% of the neurons exhibited a weak green fluorescence. Only a few noradrenergic nerve fibres were seen within the ganglia and these were associated with intraganglionic blood vessels. Small, intensely fluorescent cells were only rarely associated with the ganglia. Neurons and satellite cells formed units which were surrounded by an intraganglionic connective tissue space and a perineurium. Some of the intraganglionic capillaries were fenestrated. Neurons were entirely surrounded by satellite cells and did not appear to have any long dendrites. The generally smooth neuronal cell bodies had short spine-like processes, which were confined to within the satellite cell sheath. Preganglionic nerve fibres formed pericellular baskets of varicose fibres around neurons and made synapses either directly on the cell body or on somatic spines in about equal numbers. No synapses were found in the neuropil at a distance from the neuronal cell body. A few nerve processes were deeply embedded within the neuronal cell body. Clusters of vesicles were found in the cytoplasm of most neurons and were associated with subplasmalemmal densities. These synapse-like structures were mostly directed towards satellite cells, but some were associated with incoming synapses.

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“…The survival of ganglionic cells in the culture suggested an independence from extrapancreatic nerves. The finding is not peculiar to the pancreas; embryonic mouse heart on day 9 of gestation has been shown to have an endogenous source of cholinergic neurones which can survive in dispersed cell cultures (Lane, Sastre & Salpeter, 1976). It is postulated that in rat pancreas dissected at 14 days and grown in vitro for 7 days, there are both parasympathetic short neurones and cholinergic receptors.…”

Section: Influence Of Drugs Affecting the Autonomie Nervous System Onmentioning

confidence: 98%

Influence of Autonomic Innervation on the Foetal Rat Pancreas in Vitro

Gasparo¹,

Krinke²,

Milner³

et al. 1978

Journal of Endocrinology

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Foetal rat pancreatic rudiments, explanted on day 14 of gestation and grown for 7 days in organ culture, were highly sensitive to the stimulatory effect of glucose on insulin secretion. The autonomic innervation of the cultivated explant was investigated as a possible explanation of this phenomenon. No sign of sympathetic innervation was detected in the pancreas explanted on day 14; the uptake of alpha-methyl-noradrenaline was not detectable by fluorescence microscopy, the concentration of noradrenaline/pancreatic rudiment was less than 7 pg and no nerve-endings containing granular vesicles were visible by electron microscopy. It was concluded that sympathetic innervation of the pancreas occurs after day 14 of gestation. Sympathetic receptors developed independently of innervation since phentolamine, and, to a lesser extent, phenoxybenzamine, potentiated the gl ucose\x=req-\ induced insulin release from cultured explants. The beta-agonists isoproterenol or salbutamol were inactive alone, but in combination with phentolamine, they potentiated the effect of glucose. Alpha-antagonists reversed the inhibitory effect of adrenaline in a dose-dependent manner. Intrapancreatic neurones were demonstrated microscopically as cholinesterase\x=req-\ positive cells; parasympathetic receptors were shown to be present after potentiation of glucose-stimulated insulin release by carbamylcholine and its reversal by atropine. The preservation of parasympathetic innervation and the absence of sympathetic innervation in the cultured pancreatic rudiment may explain in part the high sensitivity of the tissue to the stimulatory effect of glucose on the release of insulin.

show abstract

Innervation of heart cells in culture by an endogenous source of cholinergic neurons.

Cited by 10 publications

References 25 publications

Phenotypic properties of adult mouse intrinsic cardiac neurons maintained in culture

Phenotypic properties of adult mouse intrinsic cardiac neurons maintained in culture

Fine structure of the autonomic ganglia of the mouse pulmonary vein

Influence of Autonomic Innervation on the Foetal Rat Pancreas in Vitro

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